Bicyclic fibrinogen antagonists

ABSTRACT

Certain compounds within formula (I) are inhibitors of platelet aggregation: ##STR1## wherein A 1  is NH or CH 2  ; 
     R is H, C 1-6  alkyl, benzyl or a carboxy protecting group; 
     R 3  is C 1-6  alkyl, Ar-C 0-6  alkyl, C 3-7  cycloalkylC 0-6  alkyl, or Het-C 0-6  alkyl; 
     R 6  is 4-amidino-Ar-N(CH 3 )CO, [[2-(4-piperidinyl)ethyl](N-methyl)amino]carbonyl, (4,4&#39;-bipiperidin-1-yl)carbonyl, [4-(2-aminoethyl)piperidin-1-yl]carbonyl, [[[3-(4-piperidinyl]propyl]methylamino]carbonyl, 1-[4-(4-pyridyl)piperazinyl]carbonyl, [[2-[(2-amino)pyrid-4-yl]ethyl]methylamino]carbonyl, [[2-(4-piperidinyl)ethyl]carbonyl]amino, [[2-(4-piperidinyl)ethyl]carbonyl]amino, [[2-(1-piperazinyl)ethyl]methylamino]-carbonyl, or [[(1,2,3,4-tetrahydro-7-isoquinolinyl]amino]carbonyl; and 
     X is H, C 1-4  alkyl, C 1-4  alkoxy, C 1-4  alkthio, trifluoroalkyl, N(R&#39;) 2 , CO 2  R&#39;, CON(R&#39;) 2 , OH, F, Cl, Br or I.

This application is a 371 of PCT/US95/00248 filed Jan. 9, 1995 which isa continuation-in-part of Ser. No. 08/179,011 filed Jan. 7, 1994, nowabandoned.

FIELD OF THE INVENTION

This invention relates to novel bicyclic compounds which inhibitplatelet aggregation, pharmaceutical compositions containing thecompounds and methods of using the compounds.

BACKGROUND OF THE INVENTION

Platelet aggregation is believed to be mediated primarily through thefibrinogen receptor, or GPIIb-IIIa platelet receptor complex, which is amember of a family of adhesion receptors referred to as integrins. Ithas been found that frequently the natural ligands of integrin receptorsare proteins which contain an Arg-Gly-Asp sequence. Von Willebrandfactor and fibrinogen, which are considered to be natural ligands forthe GPIIb-IIIa receptor, possess an Arg-Gly-Asp (RGD in single letteramino acid code) sequence in their primary structure. Functionally,these proteins are able to bind and crosslink GPIIb-IIIa receptors onadjacent platelets and thereby effect aggregation of platelets.

Fibronectin, vitronectin and thrombospondin are RGD-containing proteinswhich have also been demonstrated to bind to GPIIb-IIIa. Fibronectin isfound in plasma and as a structural protein in the intracellular matrix.Binding between the structural proteins and GPIIb-IIIa may function tocause platelets to adhere to damaged vessel walls.

Linear and cyclic peptides which bind to vitronectin and contain an RGDsequence are disclosed in WO 89/05150 (PCT US88/04403). EP 0 275 748discloses linear tetra- to hexapeptides and cyclic hexa- to octapeptideswhich bind to the GPIIb-IIIa receptor and inhibit platelet aggregation.Other linear and cyclic peptides, the disclosure of which areincorporated herein by reference, are reported in EP-A 0 341 915.However, the peptide like structures of such inhibitors often poseproblems, such as in drug delivery, metabolic stability and selectivity.Inhibitors of the fibrinogen receptor which are not constructed ofnatural amino acid sequences are disclosed in EP-A 0 372,486, EP-A 0 381033 and EP-A 0 478 363. WO 92/07568 (PCT/US91/08166) disclosesfibrinogen receptor antagonists which mimic a conformational γ-turn inthe RGD sequence by forming a monocyclic seven-membered ring structure.There remains a need, however, for novel fibrinogen receptor antagonists(e.g. inhibitors of the GPIIb-IIIa protein) which have potent in vivoand in vitro effects and lack the peptide backbone structure of aminoacid sequences.

The present invention discloses novel bicyclic compounds includingbenzazepines and benzodiazepines, which are inhibitors of the GPIIb-IIIareceptor and inhibit platelet aggregation. Certain5-phenyl-1,4-benzodiazepines are known as a class of drugs which affectthe central nervous system, and have been used as anxiolytics. SeeSternbach, L. H., J. Med. Chem., 22, 2 (1979). It has also beendisclosed that certain 5-phenyl-1,4-benzodiazepines antagonize theeffects of cholecystokinin. See Friedinger, Med. Res. Rev., 9, 271(1989). Certain bicyclic compounds which have fibrinogen antagonistactivity are disclosed in WO 93/08174 (PCT/US92/08788), WO 93/00095(PCT/US/92/05463) and WO 94/14776 (PCT/US93/12436).

SUMMARY OF THE INVENTION

In one aspect this invention is a compound as hereinafter disclosed forinhibiting platelet aggregation.

This invention is also a pharmaceutical composition for inhibitingplatelet aggregation or clot formation, which comprises a compound ofthe invention and a pharmaceutically acceptable carrier.

This invention further comprises the use of a compound as hereinafterdescribed in the manufacture of a medicament for inhibiting plateletaggregation

In another aspect, this invention provides a method for inhibitingreocclusion of an artery or vein in a mammal following fibrinolytictherapy or angioplasty, which comprises internally administering aneffective amount of a compound of the invention. This invention is alsoa method for treating stroke, transient ischemia attacks, or myocardialinfarction.

DETAILED DESCRIPTION OF THE INVENTION

Although not intending to be bound to any specific mechanism of action,the compounds of this invention are believed to inhibit the binding offibrinogen to the platelet-bound fibrinogen receptor GPIIb-IIIa, and mayinteract with other adhesion proteins via antagonism of a putative RGDbinding site.

Compounds of this invention are encompassed by formula (I): ##STR2##wherein A¹ is NH or CH₂.

R is H, C₁₋₆ alkyl, benzyl or a carboxy protecting group. Preferably Ris H.

R³ is H, C₁₋₆ alkyl, C₃₋₆ alkenyl, Ar-C₀₋₆ alkyl C₃₋₇ cycloalkylC₀₋₆alkyl, or Het-C₀₋₆ alkyl. In particular, R³ may be hydrogen, methyl,isopropyl, n-butyl, isopentyl, 2,2-dimethylbutyl, benzyl, phenylethyl,phenylpentyl, 2-thienylethyl, or cyclohexylethyl,

R⁶ is 4-amidino-Ar-N(CH₃)CO, 4-amidino-Ar-CONH,[[2-(4-piperidinyl)ethyl]methylamino]carbonyl,(4,4'-bipiperidin-1-yl)carbonyl,[4-(2-aminoethyl)piperidin-1-yl]carbonyl,[[[3-(4-piperidinyl]propyl]methylamino]carbonyl,1-[4-(4-pyridyl)piperazinyl]carbonyl,[[2-[(2-amino)pyrid-4-yl]ethyl]methylamino]carbonyl,[[2-(4-piperidinyl)ethyl]carbonyl]amino,[[2-(4-piperidinyl)ethyl]carbonyl]amino,[[2-(1-piperazinyl)ethyl]methylamino]carbonyl,[[4-(aminoiminomethyl)phenyl]carbonyl]amino, or[[(1,2,3,4-tetrahydro-7-isoquinolinyl]amino]carbonyl.

X is H, C₁₋₄ alkyl, C₁₋₄ alkoxy, C₁₋₄ alkthio, trifluoroalkyl, N(R')₂,CO₂ R', CON(R')₂, OH, F, Cl, Br or I. Preferably X is H.

Also included in this invention are pharmaceutically acceptable additionsalts, complexes or prodrugs of the compounds of this invention.Prodrugs are considered to be any covalently bonded carriers whichrelease the active parent drug according to formula (I) in vivo.

In one subgenus, A¹ is NH and forms a 1,4 benzodiazepine which ispreferably substituted in the 8 position, and R⁶ is[[2-[(2-amino)pyrid-4-yl]ethyl]methylamino]carbonyl,[[2-(4-piperidinyl)ethyl]methylamino]carbonyl, or[[4-(aminoiminomethyl)-3-fluorophenyl]methylamino]carbonyl. Preferably,R⁶ is [[2-(4-piperidinyl)ethyl]methylamino]carbonyl.

In another subgenus, A¹ is NH and forms a 1,4 benzodiazepine which issubstituted in the 7 position, and R⁶ is(4,4'-bipiperidin-1-yl)carbonyl,[[4-(aminoiminomethyl)phenyl]methylamino]carbonyl,[4-(2-aminoethyl)piperidin-1-yl]carbonyl,[[[3-(4-piperidinyl]propyl]methylamino]carbonyl,1-[4-(4-pyridyl)piperazinyl]carbonyl, or[[(1,2,3,4-tetrahydro-7-isoquinolinyl]amino]carbonyl. Preferably, R⁶ is(4,4'-bipiperidin-1-yl)carbonyl.

In yet another subgenus, A¹ is CH₂ and forms a 2-benzazepine, which issubstituted in the 7 or 8 position, and R⁶ is(4,4'-bipiperidin-1-yl)carbonyl,[[4-(aminoiminomethyl)phenyl]methylamino]carbonyl,[[2-(4-piperidinyl)ethyl]carbonyl]amino,[[2-(4-piperidinyl)ethyl]carbonyl]methylamino,[[4-(aminoiminomethyl)phenyl]carbonyl]amino or[[2-(4-piperidinyl)ethyl]methylamino]carbonyl.

Preferred compounds of this invention are: ##STR3## wherein if X is H,R³ is not phenylethyl or, with respect to (III) only, cyclohexylethyl.Such compounds wherein X is H and R³ is phenylethyl or cyclohexylethylare described in WO 94/14776 and are not a part of this invention.

General compounds of this invention include:

(R,S)-2,3,4,5-tetrahydro-4-methyl-3-oxo-8-[[[2-(4-piperidinyl)ethyl]methylamino]carbonyl]-1H-1,4-benzodiazepine-2-aceticacid;

(R,S)-2,3,4,5-tetrahydro-4-isopentyl-3-oxo-8-[[[2-(4-piperidinyl)ethyl]methylamino]carbonyl]-1H-1,4-benzodiazepine-2-aceticacid;

(R,S)-8-[[[4-(aminoiminomethyl)-3-fluorophenyl]methylamino]carbonyl]-2,3,4,5-tetrahydro-3-oxo-4-(2-phenylethyl)-1H-1,4-benzodiazepine-2-aceticacid;

(R,S)-8-[[(1,2,3,4-tetrahydro-7-isoquinolinyl]amino]carbonyl]-2,3,4,5-tetrahydro-3-oxo-4-(2-phenylethyl)-1H-1,4-benzodiazepine-2-aceticacid;

(R,S)-7-[(4,4'-bipiperidin-1-yl)carbonyl]-4-[2-(cyclohexyl)ethyl]-2,3,4,5-tetrahydro-3-oxo-1H-1,4-benzodiazepine-2-aceticacid;

(R,S)-7-[[4-(2-aminoethyl)piperidin-1-yl]carbonyl]-2,3,4,5-tetrahydro-3-oxo-4-(2-phenylethyl)-1H-1,4-benzodiazepine-2-aceticacid;

(R,S)-7-[[(1,2,3,4-tetrahydro-7-isoquinolinyl]amino]carbonyl]-2,3,4,5-tetrahydro-3-oxo-4-(2-phenylethyl)-1H-1,4-benzodiazepine-2-aceticacid;

(R,S)-7-[(4,4'-bipiperidin-1-yl)carbonyl]-2,3,4,5-tetrahydro-4-methyl-3-oxo-1H-1,4-benzodiazepine-2-aceticacid;

(R,S)-2,3,4,5-tetrahydro-3-oxo-2-(2-phenylethyl)-7-[[[2-(1-piperazinyl)ethyl]methylamino]-carbonyl]-1H-2-benzazepine-4-aceticacid;

(R,S)-2-butyl-2,3,4,5-tetrahydro-3-oxo-7-[[[2-(piperidin-4-yl)-ethyl]methylamino]carbonyl]-1H-2-benzazepine-4-aceticacid;

2-benzyl-2,3,4,5-tetrahydro-3-oxo-7-[[[2-(piperidin-4-yl)-ethyl]methylamino]carbonyl]-1H-2-benzazepine-4-acetate;

2,3,4,5-tetrahydro-3-oxo-2-(5-phenylpentyl)-7-[[[2-(piperidin-4-yl)ethyl]methylamino]carbonyl]-1H-2-benzazepine-4-acetate;

(R,S)-7-[[[4-(aminoiminomethyl)phenyl]methylamino]carbonyl]-2,3,4,5-tetrahydro-2-isopropyl-3-oxo-1H-2-benzazepine-4-aceticacid;

R,S)-2,3,4,5-tetrahydro-2-isopropyl-3-oxo-7-[[[2-(4-piperidinyl)ethyl]methylamino]carbonyl]-1H-2-benzazepine-4-aceticacid;

(R,S)-2,3,4,5-tetraydro-2-isopentyl-3-oxo-7-[[[2-(4-piperidinyl)ethyl]methylamino]carbonyl]-1H-2-benzazepine-4-aceticacid;

(R,S)-2-(3,3-dimethylbutyl)-2,3,4,5-tethydro-3-oxo-7-[[[2-(4-piperdinyl)ethyl]methylamino]carbonyl]-1H-2-benzazepine-4-aceticacid;

(R,S)-2-cyclohexyl-2,3,4,5-tetrahydro-3-oxo-7-[[[2-(4-piperidinyl)ethyl]methylamino]carbonyl]-1H-2-benzazepine-4-aceticacid;

(R,S)-2-[2-(4-fluorophenyl)ethyl]-2,3,4,5-tetrhydro-3-oxo-7-[[[2-(4-piperidinyl)ethyl]methylamino]carbonyl]-1H-2-benzazepine-4-aceticacid;

(R,S)-7-[[[4-(aminoiminomethyl)phenyl]carbonyl]amino]-2,3,4,5-tetrahydro-3-oxo-2-(2-phenylethyl]-1H-2-benzazepine-4-aceticacid;

(R,S)-2-[2-(cyclohexyl)ethyl]-2,3,4,5-tetraydro-3-oxo-7-[[[2-(4-piperidinyl)ethyl]carbonyl]amino]-1H-2-benzazepine-4-aceticacid;

(R,S)-8-[(4,4'-bipiperidin-1-yl)carbonyl]-2,3,4,5-terahydro-3-oxo-2-(2-phenylethyl)-1H-2-benzazepine-4-aceticacid;

(R,S)-2,3,4,5-tetrahydro-3-oxo-2-(2-phenylethyl)-7-[[[2-(4-piperidinyl)ethyl]methylamino]carbonyl]-1H-2-benzazepine-4-propionicacid;

(+)-7-[(4,4'-bipiperidin-1-yl)carbonyl]-2,3,4,5-tetrahydro-3-oxo-4-(phenylethyl)-1H-1,4-benzodiazepine-2-aceticacid;

(-)-7-[(4,4'-bipiperidin-1-yl)carbonyl]-2,3,4,5-tetrahydro-3-oxo-4-(phenylethyl)-1H-1,4-benzodiazepine-2-aceticacid;

(R,S)-7-[[(4,4'-bipiperidin-1-yl)]carbonyl]-2,3,4,5-tetrahydro-4-isopropyl-3-oxo-1H-1,4-benzodiazepine-2-aceticacid;

(R)-7-[(4,4'-bipiperidin-1-yl)carbonyl]-2,3,4,5-tetrahydro-4-methyl-3-oxo-1H-1,4-benzodiazepine-2-aceticacid;

(S)-7-[(4,4'-bipiperidin-1-yl)carbonyl]-2,3,4,5-tetrahydro-4-methyl-3-oxo-1H-1,4-benzodiazepine-2-aceticacid;

Sodium(R,S)-7-[(4,4'-bipiperidin-1-yl)carbonyl]-2,3,4,5-tetrahydro-4-methyl-3-oxo-1H-1,4-benzodiazepine-2-acetate;

(R)-2,3,4,5-tetrahydro-4-methyl-3-oxo-8-[[[2-(4-piperidinyl)ethyl]methylamino]carbonyl]-1H-1,4-benzodiazepine-2-aceticacid;

(S)-2,3,4,5-tetrahydro-4-methyl-3-oxo-8-[[[2-(4-piperidinyl)ethyl]methylamino]carbonyl]-1H-1,4-benzodiazepine-2-aceticacid;

(R,S)-8-[[(4,4'-bipiperidin-1-yl)]carbonyl]-2,3,4,5-tetrahydro-2-methyl-3-oxo-1H-2-benzazepine-4-aceticacid;

(R,S)-2,3,4,5-tetrahydro-7-[1-[4-(4-pyridyl)piperazinyl]carbonyl]-4methyl-3-oxo-1H-1,4-benzodiazepine-2-aceticacid;

(R,S)-7-[[[4-(aminoiminomethyl)phenyl]methylamino]carbonyl]-2,3,4,5-tetraydro-2-[2-(2-thienyl)ethyl]-3-oxo-1H-2-benzazepine-4-aceticacid;

(R,S)-2,3,4,5-tetraydro-3-oxo-4-(2-phenylethyll)-7-[[[3-(4-piperidinyl]propyl]methylamino]carbonyl]-1H-1,4-benzodiazepine-2-aceticacid;

(R,S)-2,3,4,5-tetrahydro-4-methyl-3-oxo-7-[[[3-(4-piperidnyl]propyl]methylamino]carbonyl]-1H-1,4-benzodiazepine-2-aceticacid;

(R,S)-2,3,4,5-tetydro-4-isopropyl-3-oxo-8-[[[2-(4-piperidinyl)ethyl]methylamino]carbonyl]-1H-1,4-benzodiazepine-2-aceticacid;

(S)-2,3,4,5-tetrahydro-4-(2-phenylethyl)-3-oxo-8-[[[2-(4-piperidinyl)ethyl]methylamino]carbonyl]-1H-1,4-benzodiazepine-2-aceticacid; and

(R,S)-7-[[[4-(aminoiminomethyl)phenyl]carbonyl]methylamino]-2,3,4,5-tetrahydro-3-oxo-2-(2-phenylethyl]-1H-2-benzazepine-4-aceticacid

Specific preferred compounds of this invention are

(R,S)-2,3,4,5-tetrahydro-4-methyl-3-oxo-8-[[[2-(4-piperidinyl)ethyl]methylamino]carbonyl]-1H-1,4-benzodiazepine-2-aceticacid;

(S)-2,3,4,5-tetrahydro-4-methyl-3-oxo-8-[[[2-(4-piperidinyl)ethyl]methylamino]carbonyl]-1H-1,4-benzodiazepine-2-aceticacid;

(R,S)-2,3,4,5-tetrahydro-3-oxo-4-methyl-8-[[[2-[(2-amino)pyrid-4-yl]ethyl]methylamino]carbonyl]-1H-1,4-benzodiazepine-2-aceticacid;

(R,S)-7-[(4,4'-bipiperidin-1-yl)carbonyl]-2,3,4,5-tetrahydro-4-methyl-3-oxo-1H-1,4-benzodiazepine-2-acetate;

Sodium(R,S)-7-[(4,4'-bipiperidin-1-yl)carbonyl]-2,3,4,5-tetrahydro-4-methyl-3-oxo-1H-1,4-benzodiazepine-2-acetate;

(S)-7-[(4,4'-bipiperidin-1-yl)carbonyl]-2,3,4,5-tetrahydro-4-methyl-3-oxo-1H-1,4-benzodiazepine-2-aceticacid;

Compounds of this invention wherein R³ is H or C₁₋₄ alkyl, particularlyH, methyl or isopropyl, have particularly favorable pharmacological andpharmacokinetic properties.

The meaning of any substituent at any one occurrence is independent ofits meaning, or any other substituent's meaning, at any otheroccurrence, unless specified otherwise. In cases wherein the compoundsof this invention may have one or more chiral centers, unless specified,this invention includes each unique nonracemic compound which may besynthesized and resolved by conventional techniques. In general,compounds which have the S configuration at the chiral ring carbon towhich the acetate group is attached are preferred.

Abbreviations and symbols commonly used in the peptide and chemical artsare used herein to describe the compounds of this invention. In general,the amino acid abbreviations follow the IUPAC-IUB Joint Commission onBiochemical Nomenclature as described in Eur. J. Biochem., 158, 9(1984).

C₁₋₄ alkyl as applied herein is meant to include optionally substitutedmethyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl and t-butyl. C₁₋₆alkyl additionally includes pentyl, n-pentyl, isopentyl, neopentyl andhexyl and the simple aliphatic isomers thereof. C₀₋₄ alkyl and C₀₋₆alkyl additionally indicates that no alkyl group need be present (e.g.,that a covalent bond is present).

C₃₋₆ alkenyl as applied herein means an optionally substituted alkylgroup of 3 to 6 carbons wherein a carbon-carbon single bond is replacedby a carbon-carbon double bond. C₃₋₆ alkenyl includes 1-propene,2-propene, 1-butene, 2-butene, isobutene and the several isomericpentenes and hexenes. Both cis and trans isomers are included.

Ar, or aryl, as applied herein, means phenyl or naphthyl, or phenyl ornaphthyl substituted by one to three moieties X. In particular, X may beH, C₁₋₄ alkyl, C₁₋₄ alkoxy, C₁₋₄ alkylthio, trifluoroalkyl, N(R')₂, CO₂R', CON(R')₂, OH, F, Cl, Br or I.

A substituent on a C₁₋₆ alkyl or C₃₋₆ alkenyl, such as X, may be on anycarbon atom which results in a stable structure, and is available byconventional synthetic techniques.

Het, or heterocycle, indicates an optionally substituted five or sixmembered monocyclic ring, or a nine or ten-membered bicyclic ringcontaining one to three heteroatoms chosen from the group of nitrogen,oxygen and sulfur, which are stable and available by conventionalchemical synthesis. Illustrative heterocycles are benzofuran,benzimidazole, benzopyran, benzothiophene, furan, imidazole, indoline,morpholine, piperidine, piperazine, pyrrole, pyrrolidine,tetrahydropyridine, pyridine, thiazole, thiophene, quinoline,isoquinoline, and tetra- and perhydro-quinoline and isoquinoline. Anyaccessible combination of up to three substituents, such as chosen fromX, on the Het ring that is available by chemical synthesis and is stableis within the scope of this invention.

C₃₋₇ cycloalkyl refers to an optionally substituted carbocyclic systemof three to seven carbon atoms, which may contain up to two unsaturatedcarbon-carbon bonds. Typical of C₃₋₇ cycloalkyl are cyclopropyl,cyclobutyl, cyclopentyl, cyclopentenyl, cyclohexyl, cyclohexenyl andcycloheptyl. Any combination of up to three substituents, such as chosenfrom X, on the cycloalkyl ring that is available by conventionalchemical synthesis and is stable, is within the scope of this invention.

t-Bu refers to the tertiary butyl radical, Boc refers to thet-butyloxycarbonyl radical, Fmoc refers to the fluorenylmethoxycarbonylradical, Ph refers to the phenyl radical, Cbz refers to thebenzyloxycarbonyl radical, BrZ refers to the o-bromobenzyloxycarbonylradical, ClZ refers to the o-chlorobenzyloxycarbonyl radical, Bzl refersto the benzyl radical, 4-MBzl refers to the 4-methyl benzyl radical, Merefers to methyl, Et refers to ethyl, Ac refers to acetyl, Alk refers toC₁₋₄ alkyl, Nph refers to 1- or 2-naphthyl and chex refers tocyclohexyl.

DCC refers to dicyclohexylcarbodiimide, DMAP refers to 4-(dimethylamino)pyridine, DIEA refers to diisopropylethyl amine, EDC refers toN-ethyl-N'-(dimethylaminopropyl)-carbodiimide. HOBt refers to1-hydroxybenzotriazole, THF refers to tetrahydrofuran, DMF refers todimethyl formamide, NBS refers to N-bromosuccinimide, Pd/C refers to apalladium on carbon catalyst, PPA refers to 1-propanephosphonic acidcyclic anhydride, DPPA refers to diphenylphosphoryl azide, BOP refers tobenzotriazol-1-yloxy-tris(dimethylamino)phosphonium hexafluorophosphate,HF refers to hydrofluoric acid, TEA refers to triethylamine, TFA refersto trifluoroacetic acid, PCC refers to pyridinium chlorochromate.

Compounds of this invention are prepared by treating, in any order, acompound of the structure: ##STR4## wherein A¹, X, R and R₃ are asdefined above, and R^(6') is R⁶ wherein any basic nitrogen is protected,with a reagent:

(i) to remove an amino protecting group from R^(6') ; and, if necessary,

(ii) to remove a carboxy protecting group from CO₂ R; and

(iii) form a pharmaceutically acceptable salt thereof.

Illustrative of R^(6') is R⁶ wherein the amidino, amino or piperidinylgroup is protected by a benyloxy, t-butoxycarbonyl or trifluoroacetylgroup.

The compounds of formula (IV) are generally prepared by reacting acompound of the formula (XI) with a compound of the formula (XII):##STR5## wherein A¹ and R³ are as defined for formula (I), with anyreactive functional groups protected, R is C₁₋₆ alkyl, benzyl or acarboxy protecting group, L¹ is CO₂ H or NHR', wherein R' is H or C₁₋₄alkyl, particularly methyl.

L¹ and L² are functional groups which are capable of reacting to form anamide bond. If L¹ is CO₂ H, then L² is a basic nitrogen group, such asan amino, methylamino or the nitrogen of a piperidinyl group. Forinstance R^(6") -L² is 4-amidino-Ar-NHR',2-(4-piperidinyl)(N-methyl)ethylamine, 4-(4-piperidinyl)piperidine,4-(2-aminoethyl)piperidine, 3-(4-piperidinyl)propyl(N-methyl)amine,4-(4-pyridyl)piperazine, 2-(2-aminopyrid-4-yl)ethyl(N-methyl)amine or(1,2,3,4-tetrahydro-7-isoquinolinyl)amine, wherein the non-reactivebasic nitrogen of the amidino, piperidinyl or amino group is protectedby an amino protecting group.

If L¹ is NHR', then L² is a carboxylic acid group, for instance R^(6")-L² is 3-(4-piperidinyl)propanoic acid or 4-amidino-Ar-CO₂ H, whereinthe basic nitrogen of the piperidinyl or amidino group is protected.

The compounds of formula (XI) are benzodiazepines and benzazepines, andare prepared by general methods known in the art such as those disclosedin WO 93/00095 (PCT/US/92/05463) and WO 94/14776 (PCT/US93/12436) whichare incorporated herein by reference. Compounds of formula (XII)generally contain two basic centers of which one is protected, and arelikewise known to the art. See, for instance, WO94/14776.

Coupling reagents as used herein denote reagents which may be used toform amide bonds. Typical coupling methods employ carbodiimides,activated anhydrides and esters, and acyl halides. Reagents such as EDC,DCC, DPPA, PPA, BOP reagent, HOBt, N-hydroxysuccinimide and oxalylchloride are typical.

Coupling methods to form amide bonds are generally well known to theart. The methods of peptide synthesis generally set forth by Bodansky etal., THE PRACTICE OF PEPTIDE SYNTHESIS, Springer-Verlag, Berlin, 1984,Ali et al. in J. Med. Chem., 29, 984 (1986) and J. Med. Chem., 30, 2291(1987) are generally illustrative of the technique and are incorporatedherein by reference.

Solution synthesis for the formation of amide bonds is accomplishedusing conventional methods used to form amide bonds. Typically, theamine or aniline is coupled via its free amino group to an appropriatecarboxylic acids substrate using a suitable carbodiimide coupling agent,such as N,N'-dicyclohexyl carbodiimide (DCC) or EDC, optionally in thepresence of catalysts such as 1-hydroxybenzotriazole (HOBt) anddimethylamino pyridine (DMAP). Other methods, such as the formation ofactivated esters, anhydrides or acid halides, of the free carboxyl of asuitably protected acid substrate, and subsequent reaction with the freeamine of a suitably protected amine, optionally in the presence of abase, are also suitable. For example, a protected Boc-amino acid orCbz-amidino benzoic acid is treated in an anhydrous solvent, such asmethylene chloride or tetrahydrofuran (THF), in the presence of a base,such as N-methyl morpholine, DMAP or a tralkylamine, with isobutylchloroformate to form the "activated anhydride", which is subsequentlyreacted with the free amine of a second protected amino acid or aniline.

The reactive functional groups of the sidechains of each syntheticfragment are suitably protected as known in the art. Suitable protectivegroups are disclosed in Greene, PROTECTIVE GROUPS IN ORGANIC CHEMISTRY,John Wiley and Sons, New York, 1981. For example, the Boc, Cbz,phthaloyl or Fmoc group may be used for protection of an amino (or thenitrogen of a piperidinyl) or amidino group. The Boc group is generallypreferred for protection of an amino group. A methyl, ethyl, t-Bu, cHex,benzyl, substituted benzyl, (pivaloyl)methyl or(2-methyl-2-methoxypropanoyl)methyl ester may be used for the protectionof the carboxyl group. A benzyl group or suitably substituted benzylgroup (e.g., 4-methoxy-benzyl or 2,4-dimethoxy-benzyl) is used toprotect the mercapto group or the hydroxyl group. The tosyl group may beused for protection of the imidazolyl group and tosyl or nitro group forprotection of the guanidino group. A suitably substituted carbobenzyloxygroup or benzyl group may be also be used for the hydroxyl group oramino group. Suitable substitution of the carbobenzyloxy or benzylprotecting groups is ortho and/or para substitution with chloro, bromo,nitro, methoxy or methyl, and is used to modify the reactivity of theprotective group. Except for the Boc group, the protective groups forthe amino moiety are, most conveniently, those which are not removed bymild acid treatment. These protective groups are removed by such methodsas catalytic hydrogenation, sodium in liquid ammonia or HF treatment, asknown in the art.

Methods for removal of the a carboxy or amino protecting group are wellknown in the art. For example, an alkyl or cycloalkyl ester may beremoved by basic hydrolysis, for instance an alkali metal hydroxide,such as sodium, potassium or lithium hydroxide in a suitable solvent,such as aqueous alcohol. A benzyl ester is typically removed byhydrogenation over a palladium catalyst. A basic nitrogen protected by at-butyloxycarbonyl group, or a t-butyl ester, is typically removed byacid treatment, such as by trifluoroacetic acid or hydrochloric acid,optionally diluted with a solvent, such as methylene chloride and/ordioxane. The benzyloxycarbonyl group is generally removed byhydrogenation over a palladium catalyst. A trifluoroacetyl group istypically removed by basic hydrolysis, such as by treatment with analkali metal hydroxide in a suitable solvent. One useful syntheticmethod for protecting the basic nitrogen of a piperidinyl group is tocarry the group through a synthesis as a pyridinyl group, which may bereduced with a platinum catalyst toward the end of the synthesis to"remove" the protecting group.

Acid addition salts of the compounds are prepared in a standard mannerin a suitable solvent from the parent compound and an excess of an acid,such as hydrochloric, hydrobromic, sulfuric, phosphoric, acetic, maleic,succinic or methanesulfonic. Most of the compounds form inner salts orzwitterions which may be acceptable. Cationic salts are prepared bytreating the parent compound with an excess of an alkine reagent, suchas a hydroxide, carbonate or alkoxide, containing the appropriatecation; or with an appropriate organic amine. Cations such as Li+, Na+,K+, Ca++, Mg++ and NH₄ + are specific examples of cations present inpharmaceutically acceptable salts.

This invention provides a pharmaceutical composition which comprises acompound according to formula (I) and a pharmaceutically acceptablecarrier. Accordingly, the compounds of formula (I) may be used in themanufacture of a medicament. Pharmaceutical compositions of thecompounds of formula (I) prepared as hereinbefore described may beformulated as solutions or lyophilized powders for parenteraladministration. Powders may be reconstituted by addition of a suitablediluent or other pharmaceutically acceptable carrier prior to use. Theliquid formulation may be a buffered, isotonic, aqueous solution.Examples of suitable diluents are normal isotonic saline solution,standard 5% dextrose in water or buffered sodium or ammonium acetatesolution. Such formulation is especially suitable for parenteraladministration, but may also be used for oral administration orcontained in a metered dose inhaler or nebulizer for insufflation. Itmay be desirable to add excipients such as polyvinylpyrrolidone,gelatin, hydroxy cellulose, acacia, polyethylene glycol, mannitol,sodium chloride or sodium citrate.

Alternately, these compounds may be encapsulated, tableted or preparedin a emulsion or syrup for oral administration. Pharmaceuticallyacceptable solid or liquid carriers may be added to enhance or stabilizethe composition, or to facilitate preparation of the composition. Solidcarriers include starch, lactose, calcium sulfate dihydrate, terra alba,magnesium stearate or stearic acid, talc, pectin, acacia, agar orgelatin. Liquid carriers include syrup, peanut oil, olive oil, salineand water. The carrier may also include a sustained release materialsuch as glyceryl monostearate or glyceryl distearate, alone or with awax. The amount of solid carrier varies but, preferably, will be betweenabout 20 mg to about 1 g per dosage unit. The pharmaceuticalpreparations are made following the conventional techniques of pharmacyinvolving milling, mixing, granulating, and compressing, when necessary,for tablet forms; or milling, mixing and filling for hard gelatincapsule forms. When a liquid carrier is used, the preparation will be inthe form of a syrup, elixir, emulsion or an aqueous or non-aqueoussuspension. Such a liquid formulation may be administered directly p.o.or filled into a soft gelatin capsule.

For rectal administration, the compounds of this invention may also becombined with excipients such as cocoa butter, glycerin, gelatin orpolyethylene glycols and molded into a suppository.

The compounds of this invention may be used in vitro to inhibit theaggregation of platelets in blood and blood products, e.g., for storage,or for ex vivo manipulations such as in diagnostic, therapeutic orresearch use.

This invention also provides a method of inhibiting platelet aggregationand clot formation in a mammal, especially a human, which comprises theinternal administration of a compound of formula (I) and apharmaceutically acceptable carrier. Indications for such therapyinclude acute myocardial infarction (AMI), deep vein thrombosis,pulmonary embolism, dissecting anurysm, transient ischemia attack (TIA),stroke and other infarct-related disorders, and unstable angina. Thecompounds of this invention are also useful for preventing restenosis ofan artery or vein in a mammal following angioplasty. Chronic or acutestates of hyper-aggregability, such as disseminated intravascularcoagulation (DIC), septicemia, surgical or infectious shock,post-operative and post-partum trauma, cardiopulmonary bypass surgery,incompatible blood transfusion, abruptio placenta, thromboticthrombocytopenic purpura (TTP), snake venom and immune diseases, arelikely to be responsive to such treatment. These compounds are alsobelieved to be useful for adjunct therapy following angioplasty. Inaddition, the compounds of this invention may be useful in a method forthe prevention of metastatic conditions, the prevention or treatment offungal or bacterial infection, inducing immunostimulation, treatment ofsickle cell disease, and the prevention or treatment of diseases inwhich bone resorption is a factor, such as osteoporosis.

The compounds of this invention may also be favorably administered withother agents which inhibit platelet aggregation. For instance, thecompounds of this invention may be administered with compounds of theclass of cyclooxygenase inhibitors, thromboxane antagonists, thromboxanesynthetase inhibitors, heparins, thrombin inhibitors, ADP receptorinhibitors/antagonists and ticlopidine. Examples of such agents areaspirin, warfarin and clopidogrel.

The peptide is administered either orally or parenterally to thepatient, in a manner such that the concentration of drug in the plasmais sufficient to inhibit platelet aggregation, or other such indication.The pharmaceutical composition containing the peptide Is administered ata dose between about 0.2 to about 50 mg/kg in a manner consistent withthe condition of the patient. For acute therapy, parenteraladministration is preferred. For persistent states ofhyperaggregabillty, an intravenous infusion of the peptide in 5%dextrose in water or normal saline is most effective, although anintramuscular bolus injection may be sufficient.

For chronic, but noncritical, states of platelet aggregability, oraladministration of a capsule or tablet, or a bolus intramuscularinjection is suitable. The compound is administered one to four timesdaily at a level of about 0.4 to about 50 mg/kg to achieve a total dailydose of about 0.4 to about 200 mg/kg/day.

This invention further provides a method for inhibiting the reocclusionor restenosis of an artery or vein following fibrinolytic therapy, whichcomprises internal administration of a compound of formula (I) and afibrinolytic agent. It has been found that administration of an compoundin fibrinolytic therapy either prevents reocclusion completely orprolongs the time to reocclusion.

When used in the context of this invention the term fibrinolytic agentis intended to mean any compound, whether a natural or syntheticproduct, which directly or indirectly causes the lysis of a fibrin clot.Plasminogen activators are a well known group of fibrinolytic agents.Useful plasminogen activators include, for example, anistreplase,urokinase (UK), pro-urokinase (pUK), streptokinase (SK), tissueplasminogen activator (tPA) and mutants, or variants, thereof, whichretain plasminogen activator activity, such as variants which have beenchemically modified or in which one or more amino acids have been added,deleted or substituted or in which one or more or functional domainshave been added, deleted or altered such as by combining the active siteof one plasminogen activator with the fibrin binding domain of anotherplasminogen activator or fibrin binding molecule. Other illustrativevariants include tPA molecules in which one or more glycosylation siteshave been altered. Preferred among plasminogen activators are variantsof tPA in which the primary amino acid sequence has been altered in thegrowth factor domain so as to increase the serum half-life of theplasminogen activator. tPA Growth factor variants are disclosed, e.g.,by Robinson et al., EP-A 0 297 589 and Browne et al., EP-A 0 240 334.Other variants include hybrid proteins, such as those disclosed in EP 0028 489, EP 0 155 387 and EP 0 297 882, all of which are incorporatedherein by reference. Anistreplase is a preferred hybrid protein for usein this invention. Fibrinolytic agents may be isolated from naturalsources, but are commonly produced by traditional methods of geneticengineering.

Useful formulations of tPA, SK, UK and pUK are disclosed, for example,in EP-A 0 211 592, EP-A 0 092 182 and U.S. Pat. No. 4,568,543, all ofwhich are incorporated herein by reference. Typically the fibrinolyticagent may be formulated in an aqueous, buffered, isotonic solution, suchas sodium or ammonium acetate or adipate buffered at pH 3.5 to 5.5.Additional excipients such as polyvinyl pyrrolidone, gelatin, hydroxycellulose, acacia, polyethylene, glycol, mannitol and sodium chloridemay also be added. Such a composition can be lyophilized.

The pharmaceutical composition may be formulated with both the compoundof formula (I) and fibrinolytic in the same container, but formulationin different containers is preferred. When both agents are provided insolution form they can be contained in an infusion/injection system forsimultaneous administration or in a tandem arrangement.

Indications for such therapy include myocardial infarction, deep veinthrombosis, pulmonary embolism, stroke and other infarct-relateddisorders. The compound is administered just prior to, at the same timeas, or just after parenteral administration of tPA or other fibrinolyticagent. It may prove desirable to continue treatment with the compoundfor a period of time well after reperfusion has been established tomaximally inhibit post-therapy reocclusion. The effective dose of tPA,SK, UK or pUK may be from 0.5 to 5 mg/kg and the effective dose of thepeptide may be from about 0.1 to 25 mg/kg.

For convenient administration of the inhibitor and the fibrinolyticagent at the same or different times, a kit is prepared, comprising, ina single container, such as a box, carton or other container, individualbottles, bags, vials or other containers each having an effective amountof the inhibitor for parenteral administration, as described above, andan effective amount of tPA, or other fibrinolytic agent, for parenteraladministration, as described above. Such kit can comprise, for example,both pharmaceutical agents in separate containers or the same container,optionally as lyophilized plugs, and containers of solutions forreconstitution. A variation of this is to include the solution forreconstitution and the lyophilized plug in two chambers of a singlecontainer, which can be caused to admix prior to use. With such anarrangement, the fibrinolytic and the compound may be packagedseparately, as in two containers, or lyophilized together as a powderand provided in a single container.

When both agents are provided in solution form, they can be contained inan infusion/injection system for simultaneous administration or in atandem arrangement. For example, the platelet aggregation inhibitor maybe in an i.v. injectable form, or infusion bag linked in series, viatubing, to the fibrinolytic agent in a second infusion bag. Using such asystem, a patient can receive an initial bolus-type injection orinfusion, of the inhibitor followed by an infusion of the fibrinolyticagent.

The pharmacological activity of the compounds of this invention isassessed by their ability to inhibit the binding of ³ H-SK&F 107260, aknown RGD-fibrinogen antagonist, to the GPIIbIIIa receptor; theirability to inhibit platelet aggregation, in vitro, and their ability toinhibit thrombus formation in vivo.

Inhibition of RGD-mediated GPIIb-IIIa binding

Inhibition of RGD-mediated GPIIb-IIIa binding was demonstrated byassessing the ability of compounds to inhibit the binding of ³ H-SK&F107260, a known RGD-fibrinogen antagonist, to the GPIIbIIIa receptoraccording to the procedure disclosed in WO 93/00095 (PCT/US/92/05463.

Inhibition of Platelet Aggregation

Inhibition of platelet aggregation was demonstrated following theprocedure disclosed in WO 93/00095 (PCT/US/92/05463).

The compounds of this invention generally inhibit the aggregation ofhuman platelets stimulated with ADP with IC50 of less than 0.1 uM. Thecompounds of Examples 7, 22, 23 and 26 have IC50 of between 0.2 and 11uM, and the compound of Example 4 had an IC50 of about 75 uM. Preferredcompounds have IC50 of less than 0.04 uM.

To assess the stability of the compounds to plasma proteases, thecompounds were incubated for 3 h (rather than 3 min) in the PRP prior toaddition of the agonist.

In Vivo Inhibition of Platelet Aggregation

In vivo inhibition of thrombus formation is demonstrated by recordingthe systemic and hemodynamic effects of infusion of the compounds intoanesthetized dogs according to the methods described in Aiken et al.,Prostaglandins, 19, 629 (1980). Alternately, inhibition of thrombusformation and bioavailability may be measured by the method disclosed byNichols et al., J. Pharmacol. Exp. Ther., 270, 614 (1994).

The examples which follow are intended to in no way limit the scope ofthis invention, but are provided to illustrate how to make and use thecompounds of this invention. Many other embodiments will be readilyapparent and available to those skilled in the art.

EXAMPLES

In the Examples, all temperatures are in degrees Centigrade. Massspectra were performed using fast atom bombardment (FAB) orelectro-spray (ES) ionization. Melting points were taken on aThomas-Hoover capillary melting point apparatus and are uncorrected.

NMR were recorded at 250 MHz using a Bruker AM 250 spectrometer, unlessotherwise indicated. Chemical shifts are reported in ppm (δ) downfieldfrom tetramethylsilane. Multiplicities for NMR spectra are indicated as:s=singlet, d=doutblet, t=triplet, q=quartet, m=multiplet, dd=doublet ofdoublets, dt=doublet of triplets etc. and br indicates a broad signal. Jindicates the NMR coupling constant in Hertz.

Celite® is filter aid composed of acid washed diatomaceous silica, andis a registered trademark of Mansville Corp., Denver, Colo. Florisil® isan activated magnesium silicate chromatographic support and is aregistered trademark of Floridon Co., Pittsburgh, Pa. Analtech silicagel GF and EM silica gel thin layer plates were used for thin layerchromatography. Both flash and gravity chromatography were carried outon Merck 60 (230-400 mesh) silica gel. ODS refers to an octadecylsilylderivatized silica gel chromatographic support. AN/W-TFA indicates anisocratic eluant system of the indicated percentage of acetonitrile inwater with 0.1% TFA. 5μ Apex-ODS indicates an octadecylsilanederivatized silica gel support, having a nominal particle size of 5μ,made by Jones Chromatography, Littleton, Colo. YMC ODS-AQ® is an ODSchromatographic support and is a registered trademark of YMC Co. Ltd.,Kyoto, Japan. PRP-1® is a polymeric (styrene-divinyl benzene)chromatographic support and is a registered trademark of Hamilton Co.,Reno, Nev.

Preparation 1 Preparation ofN-methyl-2-[N-(benzyloxycarbonyl)-4-piperidinyl]ethanamine

a) N-(tert-butoxycarbonyl)-N-methyl-2-(4-pyridyl)ethanamine

A solution of N-methyl-2-(4-pyridyl)ethanamine (10 g, 74 mmol) indichloromethane (250 mL) was stirred at 0° C., and treated withdi-tert-butyl dicarbonate (16 g, 74 mmol) and triethylamine (10.2 mL, 74mmol). The mixture was stirred for 24 h, diluted with chloroform andwashed with 5% sodium bicarbonate. The organic phase was dried andconcentrated to give the title compound.

b) N-(tert-butoxycarbonyl)-N-methyl-2-(4-piperidinyl)ethanamine

A solution of the compound of Preparation 1(a) (4.4 g, 18 mmol) inethanol (30 mL) was cooled to 0° C., adjusted to pH 5 with 3Nhydrochloric acid (6 mL), treated with platinum oxide (350 mg) andshaken with hydrogen for 2 h. The mixture was filtered through Celite®and the filtrate was treated with 10% sodium hydroxide (4 mL) andconcentrated to give the title compound.

c)N-(tert-butoxycarbonyl)-N-methyl-2-[N-(benzyloxycarbonyl)-4-piperidinyl]ethanamine

A mixture of the compound of Preparation 1(b) (4.4 g, 18 mmol) andtriethylamine (2.5 mL, 18 mmol) in dimethylformamide (100 mL) wasstirred at 0° C. and treated with N-(benzyloxycarbonyl)succinimide (4.4g, 18 mmol). The mixture was stirred at RT for 15 h, concentrated, andthe residue was chromatographed (silica gel, 80% ethyl acetate:hexane)to give the title compound (5.2 g, 77%).

d) N-methyl-2-[N-(benzyloxycarbonyl)-4-piperidinyl]ethanamine

A solution of the compound of Preparation 1(c) (2.2 g, 5.8 mmol) indichloromethane (15 mL) was treated with TFA (5 mL) and stirred for 2.5h. The mixture was diluted with chloroform, washed with 10% sodiumhydroxide and the organic phase was washed, dried and concentrated. Theresidue was treated with toluene and concentrated to give the titlecompound.

Preparation 2 Preparation of4-[N-(benzyloxycarbonyl)(aminoiminomethyl)]-3-fluoro-N-methyl-aniline

a) 2-fluoro-4-(methylamino)benzonitrile

To a solution of 2-fluoro-4-aminobenzonitrile [Ind. Chim. Belg., 39,490-500 (1974)] (3.8 g, 28 mmol) in anhydrous triethyl orthoformate (50mL) was added trifluoroacetic acid (0.16 g, 1.4 mmol). The solution washeated to reflux 45 min, cooled and concentrated to give a yellowcrystalline solid. To a solution of this solid in absolute ethanol (60mL) at 0° C. was added sodium borohydride (3.2 g, 84 mmol). Theresulting suspension was stirred at RT for 20 min, heated at reflux for1 h, cooled and concentrated to give a colorless solid. This residue waspartitioned between ether and water and the aqueous layer was extractedwith ether. The organic phases were combined, washed with brine anddried (sodium sulfate). Concentration gave a white solid which wasslurried in 35% ethyl acetate:hexane (200 mL) and filtered to give thetitle compound (3.7 g, 88%). mp 110-112° C.; ¹ H NMR (400 MHz, CDCl₃) δ7.35 (m, 1H), 6.4 (dd, 1H), 6.3 (dd, 1H), 2.9 (s, 3H).

b) 2-fluoro-4-(methylamino)-[N-(benzyloxycarbonyl)]benzamidine

To a suspension of ammonium chloride (3.8 g, 72 mmol) in anhydroustoluene (40 mL) at 0° C. was added a solution of trimethylaluminum intoluene (2.0 M, 72 mmol) dropwise over 5 min. The solution was stirredat RT for 45 min. The compound of Preparation 2(a) (3.6 g, 24 mmol) wasadded in one portion and the resulting solution was heated at 80° C. for22 h, cooled and poured onto a slurry of silica gel (120 g) andchloroform (350 mL). This slurry was stirred for 45 min, filtered andthe filter cake was washed with methanol (700 mL). The filtrate wasconcentrated to a yellow solid which was dissolved in a mixture of THFand water. To this solution at 0° C., 5M sodium hydroxide (14 mL, 75mmol) was added, followed by dropwise addition of benzyl chloroformate(4.1 g, 24 mmol) over 5 min. The cold solution was stirred for 30 minand concentrated. The resulting aqueous suspension was extracted withdichloromethane and then with ethyl acetate. The organic layers werecombined, dried (sodium sulfate) and concentrated to give a yellow solidwhich was purified by chromatography (silica gel, 2%methanol:dichloromethane) to give the title compound as a white powder(2.6 g, 34%). mp 128-30° C. ¹ H NMR (400 MHz, CDCl₃) δ 8.2 (t, 1H),7.5-7.25 (m, 5H), 6.4 (dd, 1H), 6.25 (dd, 1H), 5.2 (s, 2H), 4.6 (b, 1H),2.8 (d, 3H).

Preparation 3 Preparation of N²-(tert-butoxycarbonyl)-7-amino-1,2,3,4-tetrahydroisoquinoline

a) 7-nitro-1,2,3,4-tetrahydro-isoquinoline.

To a mixture of 7-nitro-1,2,3,4-tetrahydroisoquinolinone (1.50 g, 7.8mmol) in THF (15 mL) was added 2M borane methylsulfide in THF (9.0 mL,18 mmol) dropwise. After addition was complete, the clear solution washeated to reflux for 4 h. The solution was s cooled to RT, and dryhydrogen chloride was bubbled into the solution to pH 2. The solutionwas heated to reflux for 1.5 h, cooled to 0° C. and diluted with ether.The resulting white solid was filtered and dried in vacuo to yield thetitle compound (1.62 g, 76%). MS(ES) m/e 179 [M+H]⁺.

b) N² -(t-butoxycarbonyl)-7-nitro-1,2,3,4-tetraydro-isoquinoline

A solution of the compound of Preparation 3(a) (1.0 g, 5.6 mmol),di-tert-butyl dicarbonate (1.8 g, 8.4 mmol) and4-(dimethylamino)pyridine (342 mg, 2.8 mmol) in dichloromethane (50 mL)was adjusted to pH 8.0 by the addition of triethylamine. After 4 h, theresulting solution was concentrated and the residue was taken up intoethyl acetate. The solution was extracted with 5% citric acid, brine,and dried (magnesium sulfate). The organic phase was diluted withhexane, filtered and the filtrate was dried in vacuo to give the titlecompound (1.41 g, 93%). MS(ES) m/e 279 [M+H]⁺.

c) N² -(tert-butoxycarbonyl)-7-amino-1,2,3,4-tetraydro-isoquinoline

A mixture of the compound of Preparation 3(b) (1.41 g, 5.1 mmol) andPd/BaSO₄ (100 mg) in methanol (25 mL) was shaken with hydrogen (45 psi)for 1.5 h. The mixture was filtered and the filtrate concentrated toyield the title compound (1.23 g, 90% overall). MS(ES) m/e 249 [M+H]⁺.

Preparation 4 Preparation ofN-(tert-butoxycarbonyl)-2-(4-piperidinyl)ethanamine

Using the procedure of Preparation 1(a)-(b), except substituting2-(4-pyridyl)ethanamine for N-methyl-2-(4-pyridyl)ethanamine gave thetitle compound.

Preparation 5 Preparation of (5-iodopentyl)benzene

a) [5-(methanesulfonyloxy)pentyl)]benzene

Methanesulfonyl chloride (4.3 mL, 55 mmol) was added dropwise over 2 minto a solution of 5-phenylpentanol (8.4 mL, 50 mmol) and drytriethylamine (14 mL, 100 mmol) in anhydrous dichloromethane (100 mL) at0° C. under argon. After 45 min, the reaction was diluted with ether andwashed sequentially with cold 1N hydrochloric acid, water, and brine.The organic phase was dried (MgSO₄) and concentrated to give the titlecompound (12.09 g, 100%) as a pale yellow oil, which was used withoutfurther purification. ¹ H NMR (250, CDCl₃) δ 7.10-7.35 (m, 5H), 4.22 (t,J=6.5 Hz, 2H), 2.98 (s, 3H), 2.63 (t, J=7.6 Hz, 2H), 1.37-1.85 (m, 6H).

b) (5-iodopentyl)benzene

A solution of the compound of Preparation 5(a) (12.09 g, 50 mmol) andsodium iodide (8.99 g, 60 mmol) in acetone (250 mL) was heated atreflux. After 3 h, the reaction was cooled and filtered through a glassfrit, and the filtrate was concentrated. The residue was dissolved inether and washed with 1N sodium thiosulfate. The organic phase was dried(MgSO₄), concentrated, and chromatographed (silica gel, hexane) to givethe title compound (11.5 g, 84%) as a colorless oil. ¹ H NMR (250,CDCl₃) δ 7.05-7.40 (m, 5H), 3.18 (t, J=7.0 Hz, 2H), 2.62 (t, J=7.6 Hz,2H), 1.79-1.93 (m, 2H), 1.37-1.72 (m, 4H); IR (CCl₄) 3020, 2930, 2850,1495, 1451, 1200 cm⁻¹ ; MS (DCI/NH₃) m/e 292.1 [M+NH₄ ]⁺.

Preparation 6 Preparation ofN-methyl-3-[N-(tert-butoxycarbonyl)piperidin-4-yl]propanamine

The compound of Preparation 6(a) (2.0 g, 8.3 mmol) was dissolved indichloromethane (25 mL), treated with triethylamine (0.86 g, 8.6 mmol)and stirred at 0° C. The resulting mixture was treated with triflicanhydride (2.5 g, 8.9 mmol), stirred for 30 min at 0° C. and slowlyadded to a solution of methylamine in dichloromethane at 0° C. Themixture was stirred and allowed to warm to RT for 1 h. The mixture wasconcentrated and the residue was flash chromatographed (silica gel,5-20% methanol:chloroform) to give the title compound (2.8 g). MS(ES)m/e 257 [M+H⁺.

Preparation 7 Preparation of(R,S)-[7-[[4-(aminoiminomethyl)phenyl]amino]carbonyl]-4-(2-phenylethyl)-1,3,4,5-tetrahydro-3-oxo-2H-1,4-benzodiazepine-2-aceticacid

a) t-butyl 3-methyl-4-nitro-benzoate

Benzenesulfonyl chloride (12.8 mL, 100 mmol) was added rapidly to asolution of 3-methyl-4-nitrobenzoic acid (9.06 g, 50 mmol) in drypyridine (25 mL) at 40° C. The reaction was mildly exothermic. Thecloudy, orange solution was stirred for 5 min, then t-butanol (4.7 mL,50 mmol, 1 eq.) was added. After 1 h, the reddish-orange mixture waspoured into ice/water (200 mL), and the resulting mixture was stirredbriskly for 1 h. The solid was collected by suction filtration, washedwith H₂ O, and dissolved in toluene (200 mL). The solution was dried(MgSO₄) and filtered through a pad of silica gel eluting with toluene.Concentration gave the title compound as a yellow oil which crystallizedunder vacuum (9.67 g, 82%).

b) t-butyl 4-nitro-3-[N-(2-phenylethyl)aminomethyl]benzoate

A mixture of the compound of Preparation 7(a) (4.75 g, 20 mmol),N-bromosuccinimide (3.92 g, 22 mmol), and benzoyl peroxide (0.24 g, 1mmol) in CCl₄ (50 mL) was heated at reflux. After 16 h, the mixture wasfiltered to remove the succinimide, and the filtrate was concentrated toa yellow oil. The benzyl bromide was used without purification. Thecrude benzyl bromide obtained above was dissolved in dry THF (50 mL),and solid NaHCO₃ (2.52 g, 30 mmol) was added. The mixture was stirredbriskly, and phenethylamine (3.8 mL, 30 mmol) was added. The color ofthe solution darkened slightly to a deeper yellow. Within several min,the mixture had become very cloudy. After 4 h, the reaction wasconcentrated and the residue was partitioned between H₂ O (50 mL) andEt₂ O (100 mL). Separation of the phases, extraction with Et₂ O, drying(MgSO₄), and concentration gave a yellow oil. Chromatography (silicagel, 20% EtOAc/hexane) gave the title compound (2.86 g, 40%) as a yellowoil.

c) t-butyl4-nitro-3-[N-(t-butyloxycarbonyl)-N-(2-phenyl-ethyl)aminomethyl]benzoate

Di-t-butyl dicarbonate (2.10 g, 9.62 mmol) was added all at once to asolution of the compound of Preparation 7(b) (2.86 g, 8.02 mmol) inCHCl₃ (30 mL) at room temperature. The reaction was stirred at roomtemperature for 2.5 h, then at reflux for 0.5 h. Concentration andchromatography (silica gel, 15% EtOAc/hexane) gave the title compound(3.70 g,) as a yellow oil.

d) t-butyl4-amino-3-[N-(t-butyloxycarbonyl)-N-(2-phenyl-ethyl)aminomethyl]benzoate

A mixture of the compound of Preparation 7(c) (2.66 g, 5.83 mmol), 10%Pd/C (0.62 g, 0.58 mmol Pd), and EtOAc (60 mL) was shaken under H₂ (50psi). After 3 h, the mixture was filtered to remove the catalyst, andthe filtrate was concentrated to dryness. Chromatography (silica gel,20% EtOAc/hexane) gave the title compound as a yellow foamy oil whichslowly partially solidified (2.26 g, 91%).

e) t-butyl(R,S)-4-[2-(1,4-dimethoxy-1,4-dioxo-2-butyl)-amino]-3-[N-(t-butyloxycarbonyl)-N-(2-phenylethyl)-aminomethyl]-benzoate

A mixture of the compound of Preparation 7(d) (1.98 g, 4.64 mmol),dimethylacetylene dicarboxylate (1.14 mL, 9.28 mmol), and MeOH (9.3 mL)was heated to reflux under argon. A homogeneous solution resulted. After1 h, the solution was concentrated to a yellow oil. Chromatography(silica gel, 1:1 EtOAc:hexane) gave a yellow oil. This was used withoutfurther purification. TLC R_(f) 0.61 (major product), R_(f) 0.41 (minorproduct) (30% EtOAc/hexane).

The yellow oil obtained above was dissolved in EtOAc (93 mL), and 10%Pd/C (1.48 g, 1.39 mmol Pd, 0.3 eq.) was added. The mixture was shakenat RT under H₂ (45 psi) for 5 h, then was filtered to remove thecatalyst. Concentration of the filtrate gave a colorless oil which waschromatographed (silica gel, 20% EtOAc/hexane) to afford the titlecompound as a colorless oil (2.49 g, 94%).

f) methyl(R,S)-7-carboxy-4-(2-phenylethyl)-1,3,4,5-tetrahydro-3-oxo-2H-1,4-benzodiazepine-2-aceticacid

TFA (9 mL) was added all at once to a solution of the compound ofPreparation 7(e) (2.11 g, 3.70 mmol) in dry CH₂ Cl₂ (9 mL) at 0° C.under argon. The resulting light yellow solution was warmed to roomtemperature, stirred for 2 h, and concentrated. The residue wasdissolved and reconcentrated from toluene to remove residual TFA. Theresultant light yellow oil was dissolved in anhydrous MeOH (18.5 mL),and the solution was cooled to 0° C. under argon. Freshly preparedNaOMe/MeOH (1.0M; 18.5 mL, 18.5 mmol) was added dropwise over 5 min. Theice bath was removed, the yellow solution was allowed to warm to roomtemperature over 10 min, and was heated to reflux under argon. After 1.5h, the reaction was cooled in ice and quenched with glacial acetic acid(2.1 mL, 37 mmol). The mixture was concentrated and the residue waspartitioned between EtOAc (50 mL) and H₂ O (50 mL). The layers wereseparated and the aqueous layer was extracted exhaustively with EtOAc(in 50 mL portions) until all solids had dissolved. The combined organiclayers were dried (MgSO₄) and concentrated until crystallization wasinitiated. The resulting mixture was cooled thoroughly in ice. Suctionfiltration afforded the title compound as colorless crystals (1.0 g,71%). The mother liquors were concentrated and the residue waschromatographed (silica gel, 10% MeOH/CHCl₃ -trace acetic acid) toafford additional crude material. Recrystallization from EtOAccontaining a little MeOH yielded an additional amount of the pure titlecompound (1.23 g, 87%, total isolated yield).

g) methyl(R,S)-[7-[[4-[N-(benzyloxycarbonyl)amino-iminomethyl)]phenyl]aamino]carbonyl]-1,3,4,5-tetrahydro-3-oxo-4-(2-phenylethyl)-2H-1,4-benzodiazepine-2-acetate

The compound of Preparation 7(f) (95.6 mg, 0.25 mmol) was refluxed withSOCl₂ (2.5 mL) for 15 min, and the yellow solution was concentrated todryness. The yellow oily solid was dissolved in dry CH₂ Cl₂ (2.5 mL),and 4-[N-(benzyloxycarbonyl)aminoimino-methyl]aniline (134.7 mg, 0.5mmol) was added. The mixture was cooled in ice/H₂ O under argon, andanhydrous pyridine (0.061 mL, 0.75 mmol) was added dropwise. Theresulting orangish-yellow mixture was warmed to room temperature. After1.5 h, the reaction was quenched with 5% NaHCO₃ (5 mL) and extractedthoroughly with EtOAc. Drying (Na₂ SO₄), concentration, chromatography(silica gel, 3:2 EtOAc:CHCl₃ -0.5% MeOH), and preparative TLC of mixedfractions from the chromatography (same solvent system) gave the titlecompound as a pale yellow oil (110.2 mg, 70%).

h)(R,S)-[7-[[4-(aminoiminomethyl)phenyl]amino]carbonyl]-4-(2-phenylethyl)-1,3,4,5-tetrahydro-3-oxo-2H-1,4-benzodiazepine-2-aceticacid

A solution of the compound of Preparation 7(g) (87.0 mg, 0.1373 mmol) in1:1 EtOAc:MeOH (4.6 mL) containing 10% Pd/C (29.2 mg, 0.0275 mmol) andTFA (0.011 mL, 0.137 mmol) was stirred briskly under H₂. After 1 h, themixture was filtered to remove the catalyst, and the filter pad waswashed thoroughly with EtOAc and EtOAc/MeOH. Concentration of thesolution provided a yellow, oily solid (83.0 mg). This was dissolved inMeOH (4.6 mL), and 1.0N NaOH (0.41 mL, 0.41 mmol) was added. The lightyellow solution was stirred at room temperature overnight, cooled to 0°C., and acidified with TFA (0.106 mL, 1.373 mmol). The solution wasconcentrated to dryness to provide an orangish-yellow oil. This materialwas purified by preparative HPLC (PRP-1®, gradient, A:5% CH₃ CN/H₂O-0.1% TFA) for 5 min, increased to B:23% CH₃ CN/H₂ O-0.1% TFA, A for 5min, A to B during 24 min). The fractions containing the pure productwere combined and concentrated until precipitation occurred. The solidwas dissolved by addition of a minimum of CH₃ CN, and the solution waslyophillized to give the title compound as a faintly yellow powder(50%).

Preparation 8 Preparation of(R,S)-7-[[[4-(aminoiminomethyl)phenyl]-methylamino]carbonyl]-2,3,4,5-tetrahydro-3-oxo-2-(2-phenylethyl)-1H-2-benzazepine-4-aceticacid

a) t-butyl 3-bromo-4-methylbenzoate

Dimethylformamide dimethyl acetal (48 mL, 200 mmol) was added dropwiseover 15 min to a suspension of 3-bromo-4-methylbenzoic acid (85%; 10.75g, 42.5 mmol) in toluene (100 mL) at 70° C. The reaction was stirred at70-80° C. for an additional 0.5 h, then was cooled, washed sequentiallywith water and 5% sodium bicarbonate, and dried (sodium sulfate). Themixture was filtered through a pad of silica gel, and the filter pad waswashed with toluene. The filtrate was concentrated to afford the titlecompound (8.0 g, 69%) as a yellow oil. TLC (toluene) R_(f) 0.68.

b) t-butyl 3-bromo-4-(bromomethyl)benzoate

A mixture of the compound of Preparation 8(a) (1.36 g, 5 mmol),N-bromosuccinimide (0.98 g, 5.5 mmol), and benzoyl peroxide (61 mg, 0.25mmol) in carbon tetrachloride (25 mL) was heated at reflux. After 4 h,the mixture was cooled and filtered, and the filtrate was concentrated.The resulting material was used without purification. TLC R_(f) 0.39(5:95 ethyl acetate:hexane).

c) t-butyl 3-bromo-4-[[(2-phenylethyl)amino]methyl]-benzoate

The compound of Preparation 8(b) was dissolved in dry ether (25 mL), andphenethylamine (1.9 mL, 15 mmol) was added. The addition was slightlyexothermic, and the reaction became cloudy. The reaction was stirred atRT overnight, then was diluted with ether (75 mL) and was washedsequentially with 5% sodium bicarbonate and brine (25 mL). Drying(MgSO₄), concentration, and silica gel chromatography (33% ethylacetate/hexane) gave the title compound (1.26 g, 65%) as a pale yellowoil. TIC R_(f) 0.42 (30:70 ethyl acetate:hexane).

d) t-butyl3-bromo-4-[[[N-(2-phenylethyl)-N-t-butoxycarbonyl]amino]methyl]benzoate

Di-t-butyl dicarbonate (845 mg, 3.88 mmol) was added all at once to asolution of the compound of Preparation 8(c) (1.26 g, 3.23 mmol) inchloroform (16 mL) at RT. The reaction was stirred at RT for 1.5 h, thenat reflux for 0.5 h. Concentration and chromatography (silica gel, 10%ethyl acetate/hexane) gave the title compound (1.57 g, 99%) as acolorless oil which solidified in vacuo. TLC R_(f) 0.49 (10:90 ethylacetate:hexane).

e) methyl3-methoxycarbonyl-4-[[5-(t-butoxycarbonyl)-2-[[N-(2-phenylethyl)-N-t-(butoxycarbonyl)]amino]methyl]-phenyl]-2-butenoateand methyl3-methoxycarbonyl-4-[[5-(t-butoxycarbonyl)-2-[[N-(2-phenylethyl)-N-t-butoxycarbonyl)]amino]methyl]phenyl]-3-butenoate.

A mixture of the compound of Preparation 8(d) (1.34 g, 2.73 mmol),dimethyl itaconate (648 mg, 4.10 mmol), palladium (II) acetate (30.7 mg,0.14 mmol), tri-o-tolylphosphine (83.2 mg, 0.27 mmol), dry triethylamine(0.76 mL, 5.46 mmol), and dry acetonitrile (27 mL) was deoxygenatedthrough a single evacuation/argon purge cycle, then was heated at refluxunder argon. After 6 h, the reaction was cooled, and more palladium (II)acetate (30.7 mg, 0.14 mmol) and tri-o-tolylphosphine (83.2 mg, 0.27mmol) were added. The mixture was deoxygenated through threeevacuation/argon purge cycles, then was heated at reflux under argonovernight (16.5 h). The reaction was concentrated, and the residue wasdissolved in ether and washed with water and brine. Drying (MgSO₄),concentration, and chromatography (silica gel, 15% ethyl acetate/hexane;then 40% ethyl acetate/hexane) gave the crude title compound as a yellowoil. The residue was rechromatographed (silica gel, 25% ethylacetate/hexane) to yield the title compound (1.36 g, 88%) as a lightyellow oil. This material was used without separation of the isomericreaction products.

f) methyl3-methoxycarbonyl-4-[5-(t-butoxycarbonyl)-2-[[[N-(2-phenylethyl)-N-t-butoxycarbonyl]amino]methyl]-phenyl]butanoate

The compound of Preparation 8(e) (1.18 g, 2.08 mmol) was dissolved inanhydrous methanol (21 mL), and palladium (II) hydroxide on carbon (0.21g) was added. The resulting mixture was shaken at RT under H₂ (47 psi)for 2 h, then was filtered through Celite®. The filtrate wasconcentrated and resubmitted to the same reaction conditions. Afteranother 5.5 h, the mixture was filtered as before, and the filtrate wasconcentrated to afford the title compound (1.13 g, 95%) as a colorlessoil.

g) methyl(R,S)-7-carboxy-2,3,4,5-tetrahydro-3-oxo-2-(2-phenylethyl)-1H-2-benzazepine-4-acetate

TFA (11 mL) was added all at once to a cloudy solution of the compoundof Preparation 8(f) (1.30 g, 2.28 mmol) in dry CH₂ Cl₂ (11 mL) at 0° C.under argon. The resulting light yellow solution was warmed to RT,stirred for 2 h, and concentrated. The residue was concentrated oncefrom 1,2-dichloroethane to remove residual TFA and give methyl3-methoxycarbonyl-4-[5-carboxy-2-[[N-(2-phenylethyl)amino]methyl]phenyl]butanoateas a pale green oil.

The oil was dissolved in anhydrous methanol (11 mL), and the solutionwas cooled to 0° C. under argon. Freshly prepared 1.0 M sodiummethoxide/methanol (11 mL, 11 mmol) was added and the ice bath wasremoved. The yellow solution was allowed to warm to RT over 5 min andheated to reflux under argon. After 3 h, the reaction was cooled in iceand quenched with glacial acetic acid (1.3 mL, 22.8 mmol). The reactionwas diluted with ethyl acetate (100 mL) and washed with water. Thecombined aqueous layers were back-extracted with ethyl acetate, and thecombined ethyl acetate layers were washed with brine, dried (MgSO₄), andconcentrated to a pale green residue. Chromatography ((silica gel, 10%methanol/chloroform/0.1% acetic acid) gave the title compound as ayellow foam. Crystallization from methanol containing a littlechloroform gave the title compound (528.2 mg, 61%) as an off-whitesolid. mp 214-216° C.; TLC R_(f) 0.49 (10:90 methanol:chloroform).

h) methyl(R,S)-7-[[[4-[N-(benzyloxycarbonyl)-aminoiminomethyl]phenyl]methylamino]carbonyl]-2,3,4,5-tetrahydro-3-oxo-2-(2-phenylethyl)-1H-2-benzazepine-4-acetate

The compound of Preparation 8(g) (117.9 mg, 0.31 mmol) was refluxed withthionyl chloride (3 mL) for 15 min and the yellow solution wasconcentrated. The residue was concentrated from dry toluene (3 mL) toremove residual thionyl chloride, and the resulting material wasdissolved in dry CH₂ Cl₂ (0.5 mL). This solution was added dropwise over1-2 min to a solution of4-(methylamino)-(N-benzyloxycarbonyl)benzamidine (263 mg, 0.93 mmol) andanhydrous pyridine (0.125 mL, 1.55 mmol) in dry CH₂ Cl₂ (10 mL) at 0° C.under argon. The resulting yellow mixture was warmed to RT and stirredfor 0.5 h, diluted with ethyl acetate, and washed with 5% sodiumbicarbonate. Drying (sodium sulfate), concentration, and chromatography(silica gel, 10% ethyl acetate/toluene) gave the title compound (174.8mg, 87%) as a faintly yellow oil. TLC R_(f) 0.45 (9:1 toluene:ethylacetate).

i) methyl(R,S)-7-[[[4-(aminoiminomethyl)phenyl]methylamino]carbonyl]-2,3,4,5-tetrahydro-3-oxo-2-(2-phenylethyl)-1H-2-benzazepine-4-acetate

10% Palladium on carbon (58 mg, 0.054 mmol) was added carefully to asolution of the compound of Preparation 8(h) (174.8 mg, 0.27 mmol) andTFA (0.021 mL, 0.27 mmol) in ethyl acetate/methanol (1:1, 9 mL), and themixture was stirred briskly under hydrogen (balloon pressure). After 1.5h, the mixture was filtered through Celite®, and the filter pad waswashed thoroughly with ethyl acetate and methanol. Concentration gavethe title compound.

j)(R,S)-7-[[[4-(aminoiminomethyl)phenyl]methylamino]carbonyl]-2,3,4,5-tetrahydro-3-oxo-2-(2-phenylethyl)-1H-2-benzazepine-4-aceticacid

The compound of Preparation 8(i) was dissolved in methanol (9 mL), and1.0 N sodium hydroxide (0.81 mL, 0.81 mmol) was added. The solution wasstirred at RT overnight, then was concentrated. The residue wasdissolved in water/acetonitrile (3 mL), cooled to 0° C., and acidifiedwith TFA (0.21 mL, 2.7 mmol). The faintly yellow solution wasconcentrated and the residue was purified by reversed-phase flashchromatography (C-18 silica gel, 25% AN/W-TFA). Concentration andlyophilization gave the title compound (123 mg, 67%) as a colorlesspowder.

The following Examples illustrate the manner of making thepharmacologically active compounds and compositions of this invention.

Example 1 Preparation of(R,S)-2,3,4,5-tetrahydro-4-methyl-3-oxo-8-[[[2-(4-piperidinyl)ethyl]methylamino]carbonyl]-1H-1,4-benzodiazepine-2-aceticacid

a) methyl(R,S)-8-[[[2-[N-(benzyloxycarbonyl)-4-piperidinyl]ethyl]methylamino]carbonyl]-2,3,4,5-tetrahydro-4-methyl-3-oxo-1H-1,4-benzodiazepine-2-acetate

Methyl(R,S)-8-carboxy-2,3,4,5-tetrahydro-4-methyl-3-oxo-1H-1,4-benzodiazepine-2-acetate(2.0 mmol) dissolved in DMF (30 mL) was treated with EDC (2.2 mmol),1-HOBT (285 mg, 2.1 mmol) and adjusted to pH 7 with triethylamine. Themixture was treated withN-methyl-2-[N-(benzyloxycarbonyl)-4-piperidinyl]ethanamine (2.4 mmol),stirred at RT for 48 h, concentrated and the residue was purified byflash chromatography to yield the title compound. ¹ H NMR (DMSO-d₆) δ6.5-7.5 (m, 7H), 2.6-4.1 (m, 24H), 0.9-1.8 (m, 7H); MS(ES) m/e 551[M+H]⁺, [M+HCO₃ ⁻ ]⁻ 595.

b) methyl(R,S)-2,3,4,5-tetrahydro-4-methyl-3-oxo-8-[[[2-(4-piperidinyl)ethyl]methylamino]carbonyl]-1H-1,4-benzodiazepine-2-acetate

A solution of the compound of Example 1(a) (0.06 mmol) in methanol (25mL) containing 1.0M hydrogen chloride in ether (0.6 mL) was treated with10% palladium hydroxide and the mixture was shaken in a hydrogenatmosphere (40 psi) for 1 h. The mixture was filtered and concentratedto yield the title compound. MS(ES) m/e 417 [M+H]⁺.

c)(R,S)-2,3,4,5-tetrahydro-4-methyl-3-oxo-8-[[[2-(4-piperidinyl)ethyl]methylamino]carbonyl]-1H-1,4-benzodiazepine-2-aceticacid

The compound of Example 1(b) (0.26 mmol) was dissolved in methanol (9mL), and 1.0 N sodium hydroxide (0.81 mL, 0.81 mmol) was added. Thesolution was stirred at RT overnight, and concentrated. The residue wasdissolved in water/acetonitrile (3 mL), cooled to 0° C., and acidifiedwith TFA (0.21 mL, 2.7 mmol). The solution was concentrated and theresidue was purified by reversed-phase flash chromatography (C-18 silicagel, AN/W-TFA). Concentration and lyophilization gave the titlecompound. MS(ES) m/e 403 [M+H]⁺ ; ¹ H NMR (DMSO-d₆) δ 8.1-8.6 (m, 2H),6.4-7.0 (m, 3H), 5.9-6.0 (m, 1H), 5.4-5.5 (d, 1H), 5.0-5.1 (m, 1H),3.8-3.9 (d, 1H), 1.0-3.5 (m, 22H); Anal. (C₂₄ H₃₅.25 N₄ O₈.875) calcd:C, 48.91; H, 5.71; N, 9.51. found: C, 49.19; H, 5.64; N, 9.42.

Example 2 Preparation of(R,S)-2,3,4,5-tetrahydro-4-isopentyl-3-oxo-8-[[[2-(4-piperidinyl)ethyl]methylamino]carbonyl]-1H-1,4-benzodiazepine-2-aceticacid

a) methyl(R,S)-8-carboxy-2,3,4,5-tetrahydro-4-isopentyl-3-oxo-1H-1,4-benzodiazepine-2-acetate

Anhydrous isopentylamine was bubbled into anhydrous DMF (200 mL) at 0°C. for 15 min. A solution of t-butyl 4-bromomethyl-3-nitrobenzoate (5.0g, 15.8 mmol) (Int. J. Peptide. Res., 36, 31 (1990)) in DMF (10 mL) wasadded dropwise to the cold amine solution. The solution was stirred at0° for 30 min and poured into water. The mixture was extracted withethyl acetate and the combined organic layers were washed with water,dried (sodium sulfate) and concentrated to give a yellow-brown oil. Theoil was purified by chromatography over silica gel to yield t-butyl4-(isopentylamino)methyl-3-nitrobenzoate.

A solution of the t-butyl 4-(isopentylamino)methyl-3-nitrobenzoate (5.3mmol), triethylamine (2.1 g, 21 mmol) and di-t-butyl dicarbonate (3.44g, 15.8 mmol) in THF (50 mL) was stirred 24 h. The mixture wasconcentrated and the residue dissolved in ethyl acetate (200 mL). Thesolution was extracted with water, dried (sodium sulfate) andconcentrated to give a yellow oil which was purified by chromatographyover silica gel to yield t-butyl4-[N-(t-butoxycarbonyl)-N-(isopentyl)amino-methyl]-3-nitrobenzoate.

A solution of t-butyl4-[N-(t-butoxycarbonyl)-N-(isopentyl)amino-methyl]-3-nitrobenzoate (2.27mmol) in ethanol (100 mL) containing 10% palladium on carbon (0.5 g) washydrogenated (40 psi). After 30 min, the mixture was filtered andconcentrated to give t-butyl3-amino-4-[N-(t-butoxycarbonyl)-N-(isopentyl)-aminomethyl]benzoate.

Dimethyl acetylenedicarboxylate (0.34 g, 2.4 mmol) was added to asolution of t-butyl3-amino-4-[N-(t-butoxycarbonyl)-N-(isopentyl)-aminomethyl]benzoate (2.7mmol) in methanol (50 mL). The solution was heated to reflux for 1 h,treated with dimethyl acetylenedicarboxylate (0.17 g, 1.2 mmol) andheated at reflux for an additional 1 h. The mixture was cooled,filtered, concentrated and the residual yellow oil was chromatographedover ilica gel to give t-butyl(E,Z)-4-[N-(t-butoxycarbonyl)-N-(isopentyl)-aminomethyl]-3-[2-(1,4-dimethoxy-1,4-dioxo-2-butenyl)amino]-benzoate.

A solution of t-butyl(E,Z)-4-[N-(t-butoxycarbonyl)-N-(methyl)-aminomethyl]-3-[2-(1,4-dimethoxy-1,4-dioxo-2-butenyl)amino]-benzoatein methanol (50 mL) containing 10% Pd/carbon (0.5 g) was hydrogenated(45 psi) at RT. After 4 h, the suspension was filtered and concentratedto give a pale yellow oil. The residue was purified by chromatographyover silica gel to yield t-butyl4-[N-(t-butoxycarbonyl)-N-(isopentyl)amino-methyl]-3-[2-(1,4-dimethoxy-1,4-dioxo-2-butyl)amino]benzoate.

TFA (50 mL) was added to a solution of t-butyl4-[N-(t-butoxycarbonyl)-N-(isopentyl)amino-methyl]-3-[2-(1,4-dimethoxy-1,4-dioxo-2-butyl)amino]benzoate(0.3 mmol) in CH₂ Cl₂ (10 mL). The mixture was stirred for 4 h andconcentrated to yield4-[N-(isopentyl)aminomethyl]-3-[N-[2-(1,4-dimethoxy-1,4-dioxo-2-butyl)]-N-(methyl)amino]-benzoicacid.

The benzoic acid was dissolved in methanol (20 mL) and treated with 25%sodium methoxide in methanol (0.2 mL, 0.87 mmol). The mixture was heatedto 50° C. for 2 h, cooled and treated with 1M hydrogen chloride in ether(2 mL). The mixture was concentrated to give the title compound

b) methyl(R,S)-2,3,4,5-tetrahydro-4-isopentyl-3-oxo-8-[[[2-(4-pyridyl)ethyl]methylamino]carbonyl]-1H-1,4-benzodiazepine-2-acetate

The compound of Example 2(a) (2.0 mmol) dissolved in DMF (30 mL) wastreated with EDC (2.2 mmol) and 1-HOBT (285 mg, 2.1 mmol). The mixturewas treated with N-methyl-2-(4-pyridyl)ethanamine (2.4 mmol), stirred atRT for 48 h, concentrated and the residue was purified by flashchromatography to yield the title compound.

c) methyl(R,S)-2,3,4,5-tetrahydro-4-isopentyl-3-oxo-8-[[[2-(4-piperidinyl)ethyl]methylamino]carbonyl]-1H-1,4-benzodiazepine-2-acetate

A solution of the compound of Example 2(b) (0.2 mmol) and 0.6Nhydrochloric acid (0.6 mL) in methanol (30 mL) was treated with platinumoxide (5 mg) and hydrogenated (45 psi) overnight. The mixture wasfiltered and concentrated to give the title compound. MS(ES) m/e 473[M+H⁺.

d)(R,S)-2,3,4,5-tetrahydro-4-isopentyl-3-oxo-8-[[[2-(4-piperidinyl)ethyl]methylamino]carbonyl]-1H-1,4-benzodiazepine-2-aceticacid

The compound of Example 2(c) (0.27 mmol) was dissolved in methanol (9mL), and 1.0 N sodium hydroxide (0.81 mL, 0.81 mmol) was added. Thesolution was stirred at RT overnight, then was concentrated. The residuewas dissolved in water/acetonitrile (3 mL), cooled to 0° C., andacidified with TFA (0.21 mL, 2.7 mmol). The solution was concentratedand the residue was purified by reversed-phase flash chromatography toyield the title compound. MS(ES) m/e 459 [M+H]⁺.

Example 3 Preparation of(R,S)-8-[[[4-(aminoiminomethyl)-3-fluorophenyl]methylamino]carbonyl]-2,3,4,5-tetrahydro-3-oxo-4-(2-phenylethyl)-1H-1,4-benzodiazepine-2-aceticacid

a) methyl(R,S)-8-[[[4-[N-(benzyloxycarbonyl)aminoimino-methyl]-3-fluorophenyl]methylamino]carbonyl]-2,3,4,5-tetra-hydro-3-oxo-4-(2-phenylethyl)-1H-1,4-benzodiazepine-2-acetate

Using the procedure of Preparation 8(h), except substituting methyl(R,S)-8-carboxy-2,3,4,5-tetrahydro-3-oxo-4-(2-phenylethyl)-1H-1,4-benzodiazepine-2-acetatefor the compound of Preparation 8(g), and substituting2-fluoro-4-(methylamino)-[N-(benzyloxycarbonyl)]benzamidine for4-(methylamino)-(N-benzyloxycarbonyl)benzamidine, gave the titlecompound. ¹ H NMR (400 MHz, CDCl₃) δ 8.15 (t, 1H), 7.4 (d, 2H), 7.4-7.15(m, 8H), 7.1 (d, 2H), 6.9 (dd, 1H), 6.75 (dd, 1H), 6.65 (s, 1H), 6.6 (d,1H), 6.4 (d, 1H), 5.25 (d, 1H), 5.15 (s, 2H), 4.9 (q, 1H), 4.25 (d, 1H),3.7 (s, 3H), 3.65 (m, 2H), 3.55 (d, 1H), 3.45 (s, 3H), 2.95 (dd, 1H),2.7 (m, 2H), 2.6 (dd, 1H).

b)(R,S)-8-[[[4-(aminoiminomethyl)-3-fluorophenyl]methyl-amino]carbonyl]-2,3,4,5-tetrahydro-3-oxo-4-(2-phenylethyl)-1H-1,4-benzodiazepine-2-aceticacid

A solution of the compound of Example 3(a) (0.06 mmol) in methanol (25mL) containing 1.0M hydrogen chloride in ether (0.6 mL) was treated with10% palladium on carbon (0.06 g) and the mixture was shaken in ahydrogen atmosphere (40 psi) for 1 h. The mixture was filtered andconcentrated to yield the methyl ester of the title compound.

The methyl ester (0.27 mmol) was dissolved in methanol (9 mL), and 1.0 Nsodium hydroxide (0.81 mL, 0.81 mmol) was added. The solution wasstirred at RT overnight, then was concentrated. The residue wasdissolved in water/acetonitrile (3 mL), cooled to 0° C., and acidifiedwith TFA (0.21 mL, 2.7 mmol). The solution was concentrated and theresidue was purified by reversed-phase flash chromatography to yield thetitle compound. ¹ H NMR (400 MHz, CD₃ OD) δ 7.75 (t, 1H), 7.4 (d, 1H),7.25-7.05 (m, 6H), 6.8 (d, 1H), 6.7 (s, 1H), 6.5 (d, 1H), 5.35 (d, 1H),5.05 (m, 1H), 3.8 (d, 1H), 3.65 (m, 2H), 3.4 (s, 3H), 2.9 (dd, 1H), 2.7(m, 2H), 2.6 (dd, 1H). MS(ES) m/e 518.0 [M+H]⁺.

Example 4 Preparation of(R,S)-8-[[(1,2,3,4-tetrahydro-7-isoquinolinyl]amino]carbonyl]-2,3,4,5-tetrahydro-3-oxo-4-(2-phenylethyl)-1H-1,4-benzodiazepine-2-aceticacid

a) methyl(R,S)-8-[[N-(t-butoxycarbonyl)-1,2,3,4-tetrahydro-7-isoquinolinyl]amino]carbonyl]-2,3,4,5-tetrahydro-3-oxo-4-(2-phenylethyl)-1H-1,4-benzodiazepine-2-aceticacid

Methyl(R,S)-8-carboxy-1,2,4,5-tetrahydro-3-oxo-4-(2-phenylethyl)-3H-1,4-benzodiazepine-2-acetate(2.0 mmol) dissolved in DMF (30 mL) was treated with EDC (2.2 mmol) and1-HOBT (285 mg, 2.1 mmol). The mixture was treated with N²-(tert-butoxycarbonyl)-7-amino-1,2,3,4-tetrahydro-isoquinoline (2.4mmol), stirred at RT for 48 h, concentrated and the residue was purifiedby flash chromatography to yield the title compound.

b)(R,S)-8-[[(1,2,3,4-tetrahydro-7-isoquinolinyl]amino]-carbonyl]-2,3,4,5-tetraydro-3-oxo-4-(2-phenylethyl)-1H-1,4-benzodiazepine-2-aceticacid

The compound of Example 4(a) (0.56 mmol) was dissolved in a mixture ofCH₂ Cl₂ (25 mL) and TFA (5 mL). After 1 h, the mixture was concentratedto give the methyl ester of the title compound.

The methyl ester (0.27 mmol) was dissolved in methanol (9 mL), and 1.0 Nsodium hydroxide (0.81 mL, 0.81 mmol) was added. The solution wasstirred at RT overnight, then was concentrated. The residue wasdissolved in water/acetonitrile (3 mL), cooled to 0° C., and acidifiedwith TFA (0.21 mL, 2.7 mmol). The solution was concentrated and theresidue was purified by reversed-phase flash chromatography to yield thetitle compound. Anal. [C₂₉ H₃₀ N₄ O₄.0.6(C₂ HF₃ O₂).1.5(H₂ O)] calcd: C,60.55; H, 5.64; N, 9.31. found: C, 60.45; H, 5.80; N, 9.28.

Example 5 Preparation of(R,S)-7-[(4,4'-bipiperidin-1-yl)carbonyl]-4-[2-(cyclohexyl)ethyl]-2,3,4,5-tetrahydro-3-oxo-1H-1,4-benzodiazepine-2-aceticacid

a) methyl(R,S)-7-carboxy-4-[2-(cyclohexyl)ethyl]-2,3,4,5-tetrahydro-3-oxo-1H-1,4-benzodiazepine-2-acetate

A solution of methyl(R,S)-7-carboxy-2,3,4,5-tetrahydro-3-oxo-4-(2-phenylethyl)-1H-1,4-benzodiazepine-2-acetate(1.2 mmol) and 0.6N hydrochloric acid (4.0 mL) in methanol (50 mL) wastreated with platinum oxide (120 mg) and hydrogenated (45 psi) and themixture was filtered and concentrated to give the title compound.

b) methyl(R,S)-7-[(4,4'-bipiperidin-1-yl)carbonyl]-[2-(cyclohexyl)ethyl]-2,3,4,5-tetrahydro-3-oxo-4-H-1,4-benzodiazepine-2-acetate

The compound of Example 5(a) (2.0 mmol) dissolved in DMF (30 mL) wastreated with EDC (2.2 mmol) and 1-HOBT (285 mg, 2.1 mmol). This mixturewas treated with N-(t-butoxycarbonyl)-4,4'-bipiperidine (2.4 mmol),stirred at RT for 48 h, concentrated and the residue was purified byflash chromatography to yield the title compound.

c) methyl(R,S)-7-[(4,4'-bipiperidin-1-yl)carbonyl]-4-[2-(cyclohexyl)ethyl]-2,3,4,5-tetrahydro-3-oxo-1H-1,4-benzodiazepine-2-aceticacid

The compound of Example 5(b) (0.56 mmol) was dissolved in a mixture ofCH₂ Cl₂ (25 mL) and TFA (5 mL). After 1 h, the mixture was concentratedto give the title compound.

d)R,S)-7-[(4,4'-bipiperidin-1-yl)carbonyl]-4-[2-(cyclohexyl)ethyl]-2,3,4,5-tetrahydro-3-oxo-1H-1,4-benzodiazepine-2-aceticacid

The compound of Example 5(c) (0.3 mmol) was suspended in acetone (2 mL)and treated lithium hydroxide hydrate (25 mg) in water (2 mL). Themixture was stirred overnight, treated with methanol and additionallithium hydroxide hydrate (5.5 mg) was added in two portions over 9 h.The mixture was concentrated and the aqueous residue was neutralizedwith hydrochloric acid and concentrated. The residue was placed in therefrigerator overnight and filtered. The filter cake was washed withcold water, acetone and ether to yield the title compound Anal. [C₃₀ H₄₄N₄ O₄.2.5(C₂ HF₃ O₂) .2(H₂ O)] calcd: C, 49.70; H, 6.02; N, 6.62. found:C, 49.43; H, 6.02; N, 6.62.

Example 6 Preparation of(R,S)-7-[[4-(2-aminoethyl)piperidin-1-yl]carbonyl]-2,3,4,5-tetrahydro-3-oxo-4-(2-phenylethyl)-1H-1,4-benzodiazepine-2-aceticacid

a) methyl(R,S)-7-[[4-(N-t-butoxycarbonyl)(2-aminoethyl)-piperidin-1-yl]carbonyl]-2,3,4,5-tetrahydro-3-oxo-4-(2-phenylethyl)-1H-1,4-benzodiazepine-2-acetate

MethylR,S)-7-carboxy-2,3,4,5-tetrahydro-3-oxo-4-(phenylethyl)-1H-1,4-benzodiazepine-2-acetate(2.0 mmol) dissolved in DMF (30 mL) was treated with DCC (458 mg, 2.2mmol), 1-HOBT (285 mg, 2.1 mmol) and adjusted to pH 7 withtriethylamine. The mixture was treated withN-(t-butoxycarbonyl)-2-(4-piperidinyl)ethanamine (2.4 mmol), stirred atRT for 48 h, concentrated and the residue was purified by flashchromatography to yield the title compound.

b)(R,S)-7-[[4-(2-aminoethyl)piperidin-1-yl]carbonyl]-2,3,4,5-tetrahydro-3-oxo-4-(2-phenylethyl)-1H-1,4-benzodiazepine-2-aceticacid

The compound of Example 6(a) (0.56 mmol) was dissolved in a mixture ofCH₂ Cl₂ (25 mL) and TFA (5 mL). After 1 h, the mixture was concentratedto give the methyl ester of the title compound.

The methyl ester (0.27 mmol) was dissolved in methanol (9 mL), and 1.0 Nsodium hydroxide (0.81 mL, 0.81 mmol) was added. The solution wasstirred at RT overnight, then was concentrated. The residue wasdissolved in water/acetonitrile (3 mL), cooled to 0° C., and acidifiedwith TFA (0.21 mL, 2.7 mmol). The solution was concentrated and theresidue was purified by reversed-phase flash chromatography to yield thetitle compound. MS(ES) m/e 479 [M+H]⁺, 477 [M+H]⁻.

Example 7 Preparation of(R,S)-7-[[(1,2,3,4-tetrahydro-7-isoquinolinyl]amino]carbonyl]-2,3,4,5-tetrahydro-3-oxo-4-(2-phenylethyl)-1H-1,4-benzodiazepine-2-aceticacid

Using the procedure of Example 4, except substituting methyl(R,S)-7-carboxy-2,3,4,5-tetrahydro-3-oxo-4-(2-phenylethyl)-1H-1,4-benzodiazepine-2-acetatefor methyl(R,S)-8-carboxy-2,3,4,5-tetrahydro-3-oxo-4-(2-phenylethyl)-1H-1,4-benzodiazepine-2-acetate,gave the title compound. Anal. [C₂₉ H₃₀ N₄ O₄.2(C₂ HF₃ O₂).0.75(H₂ O)]calcd: C, 53.55; H, 4.36; N, 7.57. found: C, 53.80; H, 4.56; N, 7.56.

Example 8 Preparation of(R,S)-7-[(4,4'-bipiperidin-1-yl)carbonyl]-2,3,4,5-tetrahydro-4-methyl-3-oxo-1H-1,4-benzodiazepine-2-aceticacid

a) t-butyl 3-methyl-4-nitrobenzoate

3-Methyl-4-nitrobenzoic acid (500 g, 2.76 mol) was added to pyridine(1.25 L) and stirred until it dissolved. Benzenesulfonyl chloride (609g, 3.45 mol) was added rapidly while maintaining the internaltemperature <30° C. with an ice bath. A precipitate formed towards theend of the addition and pyridine (500 mL) was added to improve stirring.The mixture was stirred for 30 min and t-butanol (523 mL) was addeddropwise over 30 min at <30° C. The mixture was stirred for 2 h,dissolved in hexane:ethyl acetate (4:1; 2.5 L) and the resulting mixturewas extracted with water. The organic phase was concentrated to give alight yellow oil which crystallized on cooling to give the titlecompound (635 g, 97%). mp 56-58° C.

b) t-butyl 3-bromomethyl-4-nitrobenzoate

A solution of the compound of Example 8(a) (635 g, 2.68 mol) in carbontetrachloride (6 L) was stirred and treated with N-bromosuccinimide(476.5 g, 2.68 mol). The mixture was irradiated and heated to refluxwith 3 flood lamps positioned below the solution level. Benzoyl peroxide(6 g) was added in four portions while the mixture was heated to refluxand the last portion was added as reflux was reached after 2 h. Themixture was irradiated at reflux for 12 h, cooled, allowed to standseveral hours, filtered and concentrated to a brown oil (870.7 g). Theoil was taken up in hexane (3 L) and ether (500 mL) (boiling), filteredand the filtrate was partly concentrated by boiling. The resultingsolution was allowed to stand overnight and the off-white crystallinesolid which formed was filtered, washed with hexane and dried to givethe title compound (396.6 g, 46.8%). mp 80-83° C.; ¹ H NMR (250 MHz) δ4.5 (2H); TLC R_(f) 0.7 (silica gel, 1:9 ethyl acetate/hexane). Thefiltrate yielded a second crop on standing (73 g). mp 61-65° C.

c) t-butyl 3-(methylamino)methyl-4-nitrobenzoate

Aqueous 40% methylamine (623 mL) was added to dimethylformamide (2.5 l)stirred at -5° C. in an acetone/ice bath. A solution of the compound ofExample 8(b) (238 g, 0.753 mol) in ethyl acetate (600 mL) was addeddropwise over 1 h while maintaining the internal temperature between 0°C. and -5° C. The clear yellow reaction mixture was stirred for anadditional 30 min and was diluted with an equal volume of water. Hexanewas added, the organic phase was separated and the aqueous phase wasextracted with hexane. The combined organic extracts were washed withwater and concentrated to give the title compound (217.8 g) as a brightyellow oil.

d) t-butyl 3-[N-(t-butoxycarbonyl)-(methylamino)-methyl]-4-nitrobenzoate

A solution of the compound of Example 8(c) (0.753 mol) in ethyl acetate(1 L) was stirred and treated with di-t-butyl dicarbonate (180.8 g,0.828 mol) which was added in portions. A vigorous reaction occurredwith rapid gas evolution which ceased after the final addition of thedicarbonate. The reaction mixture was stirred for an additional 30 min,and was extracted with 4% aqueous sodium carbonate and with water. Theorganic phase was concentrated to give a light yellow oil which wasdissoved in hexane and cooled in a Dry Ice/acetone bath to initiatecrystallization. The mixture was stored in the refrigerator, filteredand dried to give the title compound (129 g, 47%) which was used in thenext step. mp 56-58° C. ¹ H NMR (CDCl₃) δ 8.05 (2H, s), 7.95 (1H, s),4.8 (2H, d), 2.9 (2H, s), 1.6 (9H, s), 1.5 (9H, s); HPLC t_(R) 27 min(Zorbax RX C₁₈, gradient, 4.6×150 mm, A:methanol B:water-0.1% TFA,50-90% methanol during 40 min, UV detection at 210 nm).

e) t-butyl 4-amino-3-[N-(t-butoxycarbonyl)-(methylamino)methyl]benzoate

A solution of the compound of Example 8(d) (129 g, 0.352 mol) in ethylacetate/methanol (1:1, 1.5 L) was treated with 10% Pd/C (40 g) moistenedwith ethyl acetate under argon in a hydrogenation bottle. The bottle waspurged with hydrogen and shaken until the theoretical amount of hydrogenwas absorbed. The mixture filtered through Celite® and the filtratecontaining the title compound was used in the next step.

f) t-butyl(E/Z)-3-[N-(t-butoxycarbonyl)(methylamino)methyl]-4-[2-(1,4-dimethoxy-1,4-dioxo-2-butenyl)amino]benzoate

A solution of the compound of Example 8(e) (118.3 g, 0.352 mol) inmethanol:ethyl acetate (1.5 L) was stirred at RT under argon anddimethyl acetylenedicarboxylate (50 g, 0.352 mol) was added dropwiseover 30 min and the mixture was heated to reflux for 16 h. The mixturewas cooled and dimethyl acetylenedicarboxylate (6.0 mL, 0.0488 mol) wasadded in one portion. The reaction mixture was heated to reflux 2.5 h.The resulting solution containing the title compound was cooled and usedin the next step.

g) t-butyl(R,S)-3-[N-(t-butoxycarbonyl)-(methylamino)-methyl]-4-[2-(1,4-dimethoxy-1,4-dioxobutyl)amino]benzoate

A solution of the compound of Example 8(f) (˜168 g, 0.352 mol) in ethylacetate:methanol (1.5 L) was added to 10% Pd/C (20 g) moistened withethyl acetate under argon in a hydrogenation bottle. The mixture wasshaken in a hydrogen atmosphere and heated to 48° C. until thetheoretical amount of hydrogen was absorbed, cooled, vented, filteredthrough Celite® and the filtrate was concentrated to give the titlecompound (173.4 g) as a yellow oil. HPLC t_(R) 18.9 min (Zorbax RX C₁₈,4.6×150 mm, 1 mL/min, gradient, A:methanol B:water-0.1% TFA, 50-90%methanol over 40 min, UV detection at 210 nm).

h)(R,S)-4-[2-(1,4-dimethoxy-1,4-dioxobutyl)amino]-3-[(methylamino)methyl]benzoate

A solution of the compound of Example 8(g) (0.348 mol) indichloromethane (500 mL) was added over 30 min to a stirred solution ofTFA (684 mL) and dichloromethane (1550 mL); the internal was between 18°C.-25° C. The solution was stirred overnight at RT and concentrated togive the title compound (254.6 g) as a brown oil. HPLC t_(R) 19 min(Zorbax RX C₁₈ column, 4.6×150 mm, gradient, 1 mL/min, A:methanolB:water-0.1% TFA, 10-60% methanol over 50 min, UV detection at 210 nm).

i) methyl(R,S)-7-carboxy-2,3,4,5-tetrahydro-4-methyl-3-oxo-1H-1,4-benzodiazepine-2-acetate

A solution of the compound of Example 8(h) [112.7 g (0.348 mol) with TFA(141.9 g)] in methanol (2 L) was cooled to between 0° C. to -5° C. andstirred under argon. Sodium methoxide in methanol (25%; 396 mL, 1.94mol) was added over 2 h, maintaining the internal temperature between 0°C. to -5° C. Following the addition, the temperature was allowed to riseslowly and the mixture stirred for 45 min. Glacial acetic acid (42 mL)was added to the reaction mixture followed by water (1.5 L). Acrystalline precipitate began to form and the pH of the mixture wasadjusted to 4.5 by careful additions of 12N hydrochloric acid. Themixture was stirred a few min, filtered and the precipitate washed withwater and vacuum dried to give the title compound (77.6 g, 76%). ¹ H NMR(250 MHz, DMSO_(d6)) δ 7.5 (2H, m), 6.5 (2H, m), 5.5 (1H, d), 5.2 (1H,m), 3.9 (1H, d), 3.6 (3H, s), 2.9 (3H, s), 2.9-2.6 (2H, m); mp 276-277°C., HPLC tR 23.2 min (Zorbax RX C18, 4.6×150 mm, 1 mL/min, gradient,A:methanol B:water-0.1% TFA, 10-60% methanol over 50 min, UV detectionat 210 nm); Anal. (C₁₄ H₁₆ N₂ O₅) calcd: C, 57.53; H, 5.52; N, 9.58.found: C, 57.27; H, 5.41; N, 9.17; mp 280-280.5° C. (dec).

j) methyl(R,S)-7-[[1'-(t-butoxycarbonyl)-4,4'-bipiperidin-1-yl]carbonyl]-2,3,4,5-tetrahydro-4-methyl-3-oxo-1H-1,4-benzodiazepine-2-acetate

Using the procedure of Example 5(b), except substituting the compound ofExample 8(i) for methyl(R,S)-7-carboxy-2,3,4,5-tetrahydro-3-oxo-4-(2-phenylethyl)-1H-1,4-benzodiazepine-2-acetate,gave the title compound.

k) methyl(R,S)-7-[(4,4'-bipiperidin-1-yl)carbonyl]-2,3,4,5-tetrahydro-4-methyl-3-oxo-1H-1,4-benzodiazepine-2-acetate

Using the procedure of Example 5(c), except substituting the compound ofExample 8(j) for the compound of Example 5(b), gave the title compound.

l)(R,S)-7-[(4,4'-bipiperidin-1-yl)carbonyl]-2,3,4,5-tetrahydro-4-methyl-3-oxo-1H-1,4-benzodiazepine-2-aceticacid

Using the procedure of Example 5(d), except substituting the compound ofExample 8(k) for the compound of Example 5(c) and substituting sodiumhydroxide for lithium hydroxide, gave the title compound. Anal. [C₂₃ H₃₂N₄ O₄.2(C₂ HF₃ O₂)].2.5(H₂ O)] calcd: C, 46.22; H, 5.46; N, 7.99. found:C, 46.33, H, 5.45; N, 7.97.

Example 9 Preparation of(R,S)-2,3,4,5-tetrahydro-3-oxo-2-(2-phenylethyl)-7-[[[2-(1-piperazinyl)ethyl]methylamino]-carbonyl]-1H-2-benzazepine-4-aceticacid

a) methyl(R,S)-7-[[[2-[4-(benzyloxycarbonyl)-1-piperazinyl)]ethyl]methylamino]carbonyl]-2,3,4,5-tetrahydro-3-oxo-2-(2-phenylethyl)-1H-2-benzazepine-4-acetate

Methyl(R,S)-7-carboxy-2,3,4,5-tetrahydro-3-oxo-2-(2-phenylethyl)-1H-2-benzazepine-4-acetate(2.0 mmol) dissolved in DMF (30 mL) was treated with DCC (458 mg, 2.2mmol), 1-HOBT (285 mg, 2.1 mmol) and adjusted to pH 7 withtriethylamine. The mixture was treated with2-[1-[4-(benzyloxycarbonyl)piperazinyl]]-N-methylethanamine (2.4 mmol),stirred at RT for 48 h, concentrated and the residue was purified byflash chromatography to yield the title compound.

b)(R,S)-2,3,4,5-tetrahydro-3-oxo-2-(2-phenylethyl)-7-[[[2-(1-piperazinyl)ethyl]methylamino]carbonyl]-1H-2-benzazepine-4-aceticacid

The compound of Example 9(a) (0.3 mmol) was dissolved in methanol (20mL), and added to palladium hydroxide (100 mg) and 3N hydrochloric acid(1 mL). The mixture was hydrogenated (40 psi) for 6 h. The reactionmixture was filtered, the catalyst washed with acetonitrile and thefiltrate concentrated.

The methyl ester (200 mg, 0.3 mmol) was dissolved in methanol (4 mL) and1N sodium hydroxide (0.5 mL), stirred, concentrated, diluted with water,acidified with 3N hydrochloric acid and concentrated. The residue waschromatographed by HPLC to yield the title compound as white solid.

Example 10 Preparation of(R,S)-2-butyl-2,3,4,5-tetrahydro-3-oxo-7-[[[2-(piperidin-4-yl)ethyl]methylamino]carbonyl]-1H-2-benzazepine-4-aceticacid

1-(3-Dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride (1.19 g,6.23 mmol) was added all at once to a solution of methyl(R,S)-7-carboxy-2,3,4,5-tetrahydro-3-oxo-1H-2-benzazepine-4-acetate(1.44 g, 5.19 mmol), benzyl alcohol (2.7 mL, 25.95 mmol),diisopropylethylamine (1.8 mL, 10.38 mmol), and4-(dimethylamino)pyridine (761 mg, 6.23 mmol) in anhydrousdimethylformamide (26 mL) at RT. The reaction was stirred for 24 h andconcentrated to leave a pale yellow oil. The oil was diluted with ethylacetate (200 mL), and the cloudy mixture was washed sequentially with 1Nhydrochloric acid and water. The combined aqueous layers were extractedwith ethyl acetate, and the combined organic layers were dried (MgSO₄)and concentrated. Chromatography over silica gel gave methyl(R,S)-7-benzyloxycarbonyl-2,3,4,5-tetrahydro-3-oxo-1H-2-benzazepine-4-acetate(1.59 g, 83%).

methyl(R,S)-7-benzyloxycarbonyl-2,3,4,5-tetrahydro-3-oxo-1H-2-benzazepine-4-acetate(220.4 mg, 0.60 mmol) was suspended in toluene (5-10 mL), and themixture was carefully concentrated to remove water and residualsolvents. The resulting solid was dissolved in dry 1:1 THF:DMF (12 mL),and 1-iodobutane (3.0 mmol) was added. Sodium hydride (60% in mineraloil, 29 mg, 0.72 mmol) was added, causing gas evolution and slightwarming. After 15 min, the reaction was cooled to 0° C. and quenchedwith saturated aqueous ammonium chloride (2 mL). The mixture was dilutedwith ether (50 mL) and washed with water. The combined aqueous layerswere extracted with ether, and the combined organic layers were dried(MgSO₄) and concentrated. Chromatography over silica gel yielded methyl(R,S)-7-benzyloxycarbonyl-2,3,4,5-tetrahydro-2-butyl-3-oxo-1H-2-benzazepine-4-acetate.

10% Pd/C (64 mg, 0.06 mmol) was added carefully to a solution of methyl(R,S)-7-benzyloxycarbonyl-2,3,4,5-tetrahydro-2-butyl-3-oxo-1H-2-benzazepine-4-acetate(0.60 mmol) in methanol (12 mL). The mixture was purged with hydrogenand stirred briskly at RT under hydrogen (balloon pressure). After 15 h,the mixture was filtered through Celite®, and the filtrate wasconcentrated to afford methyl(R,S)-7-carboxy-2,3,4,5-tetrahydro-2-butyl-3-oxo-1H-2-benzazepine-4-acetate.

1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride (90 mg, 0.47mmol) was added to a solution of methyl(R,S)-7-carboxy-2,3,4,5-tetrahydro-2-methyl-3-oxo-1H-2-benzazepine-4-acetate(0.39 mmol), N-methyl-2-(pyrid-4-yl)ethanamine (64 mg, 0.47 mmol),1-hydroxybenzotriazole (63 mg, 0.47 mmol), and diisopropylethylamine(0.14 mL, 0.78 mmol) in dry dimethylformamide (2 mL) at RT under argon.The reaction was stirred for 23 h and concentrated. The residue waspartitioned between ethyl acetate and water. The layers were separated,and the organic phase was washed with water. The combined aqueous layerswere back-extracted with ethyl acetate, the combined organic layers weredried (sodium sulfate) and concentrated to leave a yellow oil. Thecombined aqueous layers were concentrated and sodium chloride was added.The resulting mixture was exhaustively extracted with chloroform. Thecombined organic phase was dried (sodium sulfate) and concentrated togive a yellow oil, which was combined with the material from the ethylacetate phase. Chromatography over silica gel yielded methyl(R,S)-2,3,4,5-tetahydro-2-butyl-3-oxo-7-[[[2-(pyrid-4-yl)ethyl]methylamino]carbonyl]-1H-2-benzazepine-4-acetate.

Platinum oxide (4 mg) was added to a solution of methyl(R,S)-2,3,4,5-tetrahydro-2-butyl-3-oxo-7-[[[2-(pyrid-4-yl)ethyl]methylamino]carbonyl]-1H-2-benzazepine-4-acetate(0.35 mmol) and 1N hydrochloric acid (0.35 mL, 0.35 mmol) in methanol(3.5 mL), and the mixture was stirred briskly under H₂ (balloonpressure). After 7 h, the reaction was filtered through Celite® andconcentrated, and the residue was dissolved in methanol (3.5 mL).Platinum oxide (4 mg) was added, and the mixture was stirred brisklyunder H₂ (balloon pressure) for 16.5 h. Filtration through Celite® andconcentration yielded methyl(R,S)-2,3,4,5-tetrahydro-2-butyl-3-oxo-7-[[[2-(piperidin-4-yl)ethyl]methylamino]carbonyl]-1H-2-benzazepine-4-acetate.

Methyl(R,S)-2,3,4,5-tetrahydro-2-butyl-3-oxo-7-[[[2-(piperidin-4-yl)ethyl]methylamino]carbonyl]-1H-2-benzazepine-4-acetatewas dissolved in methanol (12 mL) and cooled to 0° C. 1N Sodiumhydroxide (1.05 mL, 1.05 mmol) was added dropwise, and the solution wasallowed to warm to RT. The reaction was stirred at RT for 24 h andconcentrated. The residue was concentrated from 1:1 acetonitrile:water(4 mL) to remove methanol, the residue dissolved in 1:1acetontrile:water (4 mL), and cooled in ice. TFA (0.27 mL, 3.5 mmol) wasadded and the reaction was concentrated. The residue was purified byreversed-phase flash chromatography. Concentration and lyophilizationgave the title compound. MS(ES) m/e 444 [M+H]⁺.

Examples 11-12

a) Using the procedure of Example 10 to alkylate the 2-amino group anddebenzylate the benzyl ester, except substituting benzyl bromide or(5-iodopentyl)benzene for iodobutane, and using the procedure of Example1(a)-(b), except substituting Pd/C for palladium hydroxide, yielded thefollowing compounds: 11(a). methyl2-benzyl-2,3,4,5-tetrahydro-3-oxo-7-[[[2-(piperidin-4-yl)ethyl]methylamino]carbonyl]-1H-2-benzazepine-4-acetate.MS(ES) m/e 492 [M+H]⁺. 12(a). methyl2,3,4,5-tetrahydro-3-oxo-2-(5-phenylpentyl)-7-[[[2-(piperidin-4-yl)ethyl]methylamino]carbonyl]-1H-2-benzazepine-4-acetate.MS(ES) m/e 548 [M+H]⁺.

b) Using the procedure of Example 1(c), except substituting thecompounds of Example 11-12(a) for the compound of Example 1(b), gave thefollowing compounds: 11(b).2-benzyl-2,3,4,5-tetrahydro-3-oxo-7-[[[2-(piperidin-4-yl)ethyl]methylamino]carbonyl]-1H-2-benzazepine-4-acetate.12(b).2,3,4,5-tetrahydro-3-oxo-2-(5-phenylpentyl)-7-[[[2-(piperidin-4-yl)ethyl]methylamino]carbonyl]-1H-2-benzazepine-4-acetate.

Example 13 Preparation of(R,S)-7-[[[4-(aminoiminomethyl)phenyl]methyl-amino]carbonyl]-2,3,4,5-tetrahydro-2-isopropyl-3-oxo-1H-2-benzazepine-4-aceticacid

Using the procedure of Preparation 8(c)-(j), except substitutingisopropylamine for 2-phenethylamine in the procedure of Preparation 8(c)and substituting 2,4,6-collidine for sodium methoxide in methanol in theprocedure of Preparation 8(g), gave the title compound. MS(ES) m/e 437[M+H]⁺.

Examples 14-17

Using the procedures of Preparation 8(c)-(g), except substituting a)isopropylamine, b) isopentylamine, c) 3,3-dimethylbutylamine or d)cyclohexylamine for phenethylamine in the procedure of Preparation 8(c),and optionally substituting 2,4,6-collidine, triethylamine in toluene,or diisopropylethylamine in xylene, for sodium methoxide in methanol inthe procedure of Preparation 8(g), and subsequently using the procedureof Example 2(b)-(d), except optionally substitutingbenzotriazol-1-yloxy-tris(dimethylamino)-phosphonium hexafluorophosphatefor 1-(3-dimethylamino-propyl)-3-ethylcarbodiimide, gave the followingcompounds: 14.(R,S)-2,3,4,5-tetrahydro-2-isopropyl-3-oxo-7-[[[2-(4-piperidinyl)ethyl]methylamino]carbonyl]-1H-2-benzazepine-4-aceticacid. MS(ES) m/e 430 [M+H]⁺. 15.(R,S)-2,3,4,5-tetrahydro-2-isopentyl-3-oxo-7-[[[2-(4-piperidinyl)ethyl]methylamino]carbonyl]-1H-2-benzazepineacetic acid. MS(ES) m/e 458 [M+H]⁺. 16.(R,S)-2-(3,3-dimethylbutyl)-2,3,4,5-tetrahydro-3-oxo-7-[[[2-(4-piperidinyl)ethyl]methylamino]carbonyl]-1H-2-benzazepine-4-aceticacid. MS(ES) m/e 472 [M+H]⁺. 17.(R,S)-2-cyclohexyl-2,3,4,5-tetrahydro-3-oxo-7-[[[2-(4-piperidinyl)ethyl]methylamino]carbonyl]1-H-2-benzazepine-4-aceticacid. MS(ES) m/e 470 [M+H]⁺.

Example 18 Preparation of(R,S)-2-[2-(4-fluorophenyl)ethyl]-2,3,4,5-tetrahydro-3-oxo-7-[[[2-(4-piperidinyl)ethyl]methylamino]-carbonyl]-1H-2-benzazepine-4-aceticacid

Using the procedure of Preparation 8(c)-(g), except substituting4-(fluoro)phenethylamine for phenethylamine in the procedure ofPreparation 8(c) and substituting diisopropylethylamine in xylene forsodium methoxide in methanol in the procedure of Preparation 8(g), andusing the procedure of Example 1, gave the title compound. MS(ES) m/e510 [M+H]⁺.

Example 19 Preparation of(R,S)-7-[[[4-(aminoiminomethyl)phenyl]-carbonyl]amino]-2,3,4,5-tetrahydro-3-oxo-2-(2-phenylethyl]1-H-2-benzazepine-4-aceticacid

a) methyl(R,S)-7-amino-2,3,4,5-tetrahydro-3-oxo-2-(2-phenylethyl]-1H-2-benzazepine-4-acetate

A mixture of methyl(R,S)-7-carboxy-2,3,4,5-tetrahydro-3-oxo-2-(2-phenylethyl)-1H-2-benzazepine-4-acetate(2.67 mmol), triethylamine (0.42 mL, 5.87 mmol) and diphenylphosphorylazide (0.62 mL, 2.80 mmol) in toluene (20 mL) was heated at105° C. for 0.5 h. After the temperature was lowered to 80° C., themixture was treated with benzyl alcohol (0.60 mL, 0.42 mmol), stirredfor 14 h and concentrated. The residue was purified by flashchromatography to give methyl(R,S)-7-(carbobenzyloxy)amino-2,3,4,5-tetrahydro-3-oxo-2-(2-phenylethyl)-1H-2-benzazepine-4-acetate.

The Cbz compound (0.68 g, 1.38 mmol) was dissolved in methanol (10 mL)and palladium on carbon (5%, 50 mg) was added. The mixture washydrogenated for 1 h, filtered through Celite® and concentrated to yieldthe title compound.

b) methyl(R,S)-7-[[[4-(N-benzyloxycarbonyl)-(aminoimiomethyl)phenyl]carbonyl]amino]-2,3,4,5-tetrahydro-3-oxo-2-(2-phenylethyl]-1H-2-benzazepine-4-acetate

4-[N-(benzyloxycarbonyl)(aminoiminomethyl)]benzoic acid (2.0 mmol)dissolved in DMF (30 mL) was treated with1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride (2.2 mmol),1-HOBT (285 mg, 2.1 mmol) and adjusted to pH 7 withdiisopropylethylamine. The mixture was treated with the compound ofExample 19(a) (2.4 mmol), stirred at RT for 48 h, concentrated and theresidue was purified by flash chromatography to yield the titlecompound.

c) methyl(R,S)-7-[[[4-(aminoimiomethyl)phenyl]carbonyl]-amino]-2,3,4,5-tetrahydro-3-oxo-2-(2-phenylethyl]-1H-2-benzazepine-4-acetate

A solution of the compound of Example 19(b) (0.15 mmol) and 10% Pd/C (20mg) in methanol (40 mL) and acetic acid (8 drops) was shaken in ahydrogen atmosphere (45 psi) for 30 min. The mixture was filtered andthe filtrate concentrated in vacuo to yield the title compound.

d) methyl(R,S)-7-[[[4-(aminoiminomethyl)phenyl]carbonyl]-amino]-2,3,4,5-tetrahydro-3-oxo-2-(2-phenylethyl]-1H-2-benzazepine-4-acetate

A solution of the compound of Example 19(c) (65 mg, 0.12 mmol) wasstirred in lithium hydroxide in aqueous THF at room temperature underargon overnight. The mixture was treated with 3N HCl (1 mL) andconcentrated in vacuo. The residue was purified by HPLC. MS(ES) m/e 485[M+H]⁺, 483 {M-H]⁻.

Example 20 Preparation of(R,S)-2-[2-(cyclohexyl)ethyl]-2,3,4,5-tetrahydro-3-oxo-7-[[[2-(4-piperidinyl)ethyl]carbonyl]amino]-1H-2-benzazepine-4-aceticacid

a) methyl(R,S)-2,3,4,5-tetrahydro-3-oxo-7-[[[2-(4-pyridyl)ethyl]carbonyl]amino]-2-(phenylethyl)-1H-2-benzazepine-4-acetate

Using the procedure of Example 19(b), except substituting4-(pyridyl)propionic acid for4-[N-(benzyloxycarbonyl)(aminoiminomethyl)]benzoic acid, gave the titlecompound.

b)(R,S)-2-[2-(cyclohexyl)ethyl]-2,3,4,5-tetrahydro-3-oxo-7-[[[2-(4-piperidinyl)ethyl]carbonyl]amino]-1H-2-benzazepine-4-acetate

Using the procedure of Example 2(c), except substituting the compound ofExample 20(a) for the compound of Example 2(b) and subsequently usingthe procedure of Example 2(d) except substituting lithium hydroxide inaqueous THF for sodium hydroxide in aqueous methanol, gave the titlecompound. MS(ES) m/e 484 [M+H]⁺, 482 [M-H]⁻.

Example 21 Preparation of(R,S)-8-[(4,4'-bipiperidin-1-yl)carbonyl]-2,3,4,5-tetrahydro-3-oxo-2-(2-phenylethyl)-1H-2-benzazepine-4-aceticacid

a) methyl(R,S)-8-carboxy-2,3,4,5-tetrahydro-3-oxo-2-(2-phenylethyl)-1H-1,4-benzodiazepine-4-acetate

Using the procedure of Preparation 8(a)-(g), except substituting4-bromo-3-methylbenzoic acid for 3-bromo-4-methylbenzoic acid, gave thetitle compound.

b)(R,S)-8-[(4,4'-bipiperidin-1-yl)carbonyl]-2,3,4,5-tetrahydro-3-oxo-4-(2-phenylethyl)-1H-2-benzodiazepine-4-aceticacid

Using the procedure of Example 5(b)-(d), except substituting thecompound of Example 21(a) for the compound of Example 5(a) gave thetitle compound. Anal. (C₃₁ H₃₉ N₃ O₄.H₂ O) calcd: C, 69.78; H, 7.71; N,7.84. found: C, 69.38; H, 7.68; N, 7.37.

Example 22 Preparation of(R,S)-2,3,4,5-tetrahydro-3-oxo-2-(2-phenylethyl)-7-[[[2-(4-piperidinyl)ethyl]-methylamino]carbonyl]-1H-2-benzazepine-4-propionicacid

a) ethyl5-[5-(t-butoxycarbonyl)-2-[[[N-(t-butoxy-carbonyl)-N-(2-phenylethyl)]amino]methyl]phenyl]-4-ethoxycarbonyl-4-pentenoate;and ethyl5-[5-(t-butoxycarbonyl)-2-[[[N-(t-butoxycarbonyl)-N-(2-phenylethyl))]amino]methyl]phenyl]-4-ethoxycarbonyl-3-pentenoate.

Using the procedure of Preparation 8(e), except sub-stituting diethyl2-methyleneglutarate [Tet. Lett., 30, 7381 (1989)] for dimethylitaconate, gave the title compounds.

b) ethyl5-[5-(t-butoxycarbonyl)-2-[[[N-(t-butoxycarbonyl)-N-(2-phenylethyl)]amino]methyl]phenyl]-4-(ethoxycarbonyl)-pentanoate

Using the procedure of Preparation 8(f), except substituting thecompound of Example 22(a) for the compound of Preparation 8(e) andsubstituting Pd/C in ethanol:ethyl acetate for palladium hydroxide inmethanol, gave the title compound.

c) ethyl(R,S)-7-carboxy-2,3,4,5-tetrahydro-3-oxo-2-(2-phenylethyl)-1H-2-benzazepine-4-propanoate

Using the procedure of Preparation 8(g), except substituting thecompound of Example 22(b) for the compound of Preparation 8(f) andsubstituting hydrogen chloride in dioxane for TFA, gave the titlecompound.

d)(R,S)-2,3,4,5-tetrahydro-3-oxo-2-(2-phenylethyl)-7-[[[2-(4-piperidinyl)ethyl]methylamino]carbonyl]-1H-2-benzazepine-4-propionicacid

Using the procedure of Example 1, except substituting the compound ofExample 22(c) for methyl(R,S)-8-carboxy-2,3,4,5-tetrahydro-4-methyl-3-oxo-1H-1,4-benzodiazepine-2-acetate,gave the title compound. MS(ES) m/e 506 [M+H]⁺, 540 [M-H]⁻ ; Anal. (C₃₀H₃₉ N₃ O₄.HCl.3/8H₂ O) calcd: C, 65.68; H, 7.48; N, 7.66. found: C,65.67; H, 7.55.

Example 23-24

a) Methyl(R,S)-7-[1'-(t-butoxycarbonyl)-4,4'-bipiperidin-1-yl]carbonyl]-2,3,4,5-tetrahydro-3-oxo-4-(phenylethyl)-1H-1,4-benzodiazepine-2-acetatewas chromatographed HPLC t_(R) 16 min and 19.8 min (Chiracel® OD,21.1×250 mm, 10 mL/min, methanol, UV detection at 250 nm) to give thefollowing compounds: 23(a). methyl(+)-7-[1'-(t-butoxycarbonyl)-4,4'-bipiperidin-1-yl]carbonyl]-2,3,4,5-tetrahydro-3-oxo-4-(phenylethyl)-1H-1,4-benzodiazepine-2-acetate.24(a). methyl(-)-7-[1'-(t-butoxycarbonyl)-4,4'-bipiperidin-1-yl]carbonyl]-2,3,4,5-tetrahydro-3-oxo-4-(phenylethyl)-1H-1,4-benzodiazepine-2-acetate.

b) Using the procedure of Example 5(c)-(d), except substituting thecompounds of Example 23-24(a) for the compound of Example 5(b), gave thefollowing compounds: 23(b).(+)-7-[(4,4'-bipiperidin-1-yl)carbonyl]-2,3,4,5-tetrahydro-3-oxo-4-(phenylethyl)-1H-1,4-benzodiazepine-2-aceticacid: [α]_(D) +128 (c 0.5,CH₃₀ H); [α]D +80 (c=0.5 water); Anal. (C₃₀H₃₈ N₄ O₄.2C₂ HF₃ O₂.1.5H₂ O) calcd: C, 52.78, H, 5.60, N, 7.24. found:C, 52.70, H, 5.89, N, 7.47. 24(b).(-)-7-[(4,4'-bipiperidin-1-yl)carbonyl]-2,3,4,5-tetrahydro-3-oxo-4-(phenylethyl)-1H-1,4-benzodiazepine-2-aceticacid: [α]_(D) -118 (c 0.5,CH₃ OH); Anal. (C₃₀ H₃₈ N₄ O₄.2C₂ HF₃ O₂)calcd: C, 54.69, H, 5.40, N, 7.50. found: C, 54.49, H, 5.40, N, 7.85.

Example 25 Preparation of(R,S)-7-[[(4,4'-bipiperidin-1-yl)]carbonyl]-2,3,4,5-tetrahydro-4-isopropyl-3-oxo-1H-1,4-benzodiazepine-2-aceticacid

a) methyl(R,S)-7-carboxy-4-isopropyl-2,3,4,5-tetrahydro-3-oxo-1H-1,4-benzodiazepine-2-acetate

Using the procedure of Preparation 7(b)-(f), except substitutingisopropylamine for phenethylamine in Preparation 7(b) and substitutingdiisopropylethylamine in refluxing xylene for sodium methoxide inmethanol in Preparation 7(f), gave the title compound.

b) methyl(R,S)-7-[[(4,4'-bipiperidin-1-yl)]carbonyl]-2,3,4,5-tetrahydro-4-isopropyl-3-oxo-1H-1,4-benzodiazepine-2-acetate

Using the procedure of Example 5(b)-(c), except substituting thecompound of Example 25(a) for the compound of Example 5(a) gave thetitle compound.

c)(R,S)-7-[[(4,4'-bipiperidin-1-yl)]carbonyl]-2,3,4,5-tetrahydro-4-isopropyl-3-oxo-1H-1,4-benzodiazepine-2-aceticacid

Using the procedure of Preparation 8(j), except substituting thecompound of Example 25(b) for the compound of Preparation 8(i), gave thetitle compound. HPLC k' 4.27 (PRP-1; 15% acetonitrile/water/0.1% TFA); ¹H NMR (400 MHz, CD₃ OD) δ 7.12 (d, J=1.9 Hz, 1H), 7.08 (dd, J=8.3, 1.9Hz, 1H), 6.59 (d, J=8.3 Hz, 1H), 5.29 (d, J=17.0 Hz, 1H), 5.17 (dd,J=8.9, 5.2 Hz, 1H), 4.72-4.84 (m, 1H), 4.10 (d, J=17.0 Hz, 1H),3.34-3.45 (m, 2H), 2.70-3.15 (m, 5H), 2.63 (dd, J=16.8, 5.2 Hz, 1H),1.92-2.05 (m, 2H), 1.69-1.88 (m, 2H), 1.35-1.54 (m, 4H), 1.16-1.31 (m,2H), 1.20 (d, J=6.8 Hz, 3H), 1.03 (d, J=6.9 Hz, 3H); MS(ES) m/e 457.2[M+H]⁺ ; Anal. (C₂₅ H₃₆ N₄ O₄.1.5 CF₃ CO₂ H.1.5 H₂ O) calcd: C, 51.37;H, 6.24; N, 8.56. found: C, 51.49; H, 6.39; N, 8.46.

Examples 26-27 Preparation of(R)-7-[(4,4'-bipiperidin-1-yl)carbonyl]-2,3,4,5-tetrahydro-4-methyl-3-oxo-1H-1,4-benzodiazepine-2-aceticacid; and(S)-7-[(4,4'-bipiperidin-1-yl)carbonyl]-2,3,4,5-tetrahydro-4-methyl-3-oxo-1H-1,4-benzodiazepine-2-aceticacid

a) The compound of Example 8(j) was separated by chiral HPLC to give thefollowing compounds: 26(a). methyl(R)-7-[[1'-(t-butoxycarbonyl)-4,4'-bipiperidin-1-yl]carbonyl]-2,3,4,5-tetrahydro-4-methyl-3-oxo-1H-1,4-benzodiazepine-2-acetate.[α]_(D) +140 (c 1,CH₃ OH); HPLC t_(R) 12.9 min (Chiralcel OD, 21.2×250mm, 10 mL/min, methanol, UV detection at 325 nm) 27(b). methyl(S)-7-[[1'-(t-butoxycarbonyl)-4,4'-bipiperidin-1-yl]carbonyl]-2,3,4,5-tetrahydro-4-methyl-3-oxo-1H-1,4-benzodiazepine-2-acetate.[α]_(D) -142.5 (c 1,CH₃ OH); HPLC t_(R) 15.1 min (Chiralcel OD, 21.2×250mm, 10 mL/min, methanol, UV detection at 325 nm).

b) Using the procedure of Example 8(k), except substituting thecompounds of Example 26(a) and 27(a) for the compound of Example 8(j)gave the following compounds: 26(b). methyl(R)-7-[(4,4'-bipiperidin-1-yl)carbonyl]-2,3,4,5-tetrahydro-4-methyl-3-oxo-1H-1,4-benzodiazepine-2-acetate.HPLC t_(R) 14.3 min (Ultrasphere C18, 4.6×250 mm, 1.5 mL/min, gradient,A:acetonitrile B: water-0.1% TFA, 5-60% acetonitrile over 20 min, UVdetection at 220 nm) 27(b). methyl(S)-7-[(4,4'-bipiperidin-1-yl)carbonyl]-2,3,4,5-tetrahydro-4-methyl-3-oxo-1H-1,4-benzodiazepine-2-acetate.HPLC t_(R) 14.61 min (Ultrasphere C18, 4.6×250 mm, 1.5 mL/min, gradient,A:acetonitrile B: water-0.1% TFA, 5-60% acetonitrile over 20 min, UVdetection at 220 nm).

c) Using the procedure of Preparation 8(j), except substituting thecompounds of Example 26(b) and 27(b) for the compound of Preparation8(i) gave the following compounds: 26(c).(R)-7-[(4,4'-bipiperidin-1-yl)carbonyl]-2,3,4,5-tetrahydro-4-methyl-3-oxo-1H-1,4-benzodiazepine-2-acetate:[α]_(D) +92.4 (c 0.5,CH₃ OH); MS(ES) m/e 429 [M+H]⁺ ; 427 [M-H]⁻ ; HPLCt_(R) 12.7 min (Ultrasphere C18, 4.6×250 mm, 1.5 mL/min, gradient,A:acetonitrile B: water-0.1% TFA, 5-60% acetonitrile over 20 min, UVdetection at 220 nm); Anal. (C₂₃ H₃₂ N₄ O₄.4.5 CF₃ CO₂ H.3.33 H₂ O)calcd: C, 37.44; H, 4.16; N, 5.29. found: C, 37.43; H, 3.81; N, 5.20.27(c).(S)-7-[(4,4'-bipiperidin-1-yl)carbonyl]-2,3,4,5-tetrahydro-4-methyl-3-oxo-1H-1,4-benzodiazepine-2-acetate:[α]_(D) -105.9 (c 0.6,CH₃ OH); MS(ES) m/e 429 [M+H]⁺ ; 427 [M-H]⁻ ; HPLCt_(R) 12.6 min (Ultrasphere C18, 4.6×250 mm, 1.5 mL/min, gradient,A:acetonitrile B: water-0.1% TFA, 5-60% acetonitrile over 20 min, UVdetection at 220 nm); Anal. (C₂₃ H₃₂ N₄ O₄.3 CF₃ CO₂ H.1.25 H₂ O) calcd:C, 43.92; H, 4.77; N, 7.06. found: C, 44.09; H, 5.00; N, 7.29.

Example 28 Preparation of(S)-7-[(4,4'-bipiperidin-1-yl)carbonyl]-2,3,4,5-tetrahydro-4-methyl-3-oxo-1H-1,4-benzodiazepine-2-aceticacid

a) 4-fluoro-3-methylbenzoic acid

A solution of 4-fluoro-3-methylphenylmagnesium bromide in THF (1.0 M,250 mL, 250 mmol) was added in a stream over 5 min to a mixture of DryIce (10 g) in dry THF (250 mL), and the reaction was allowed to warm toRT and concentrated. The residue was partitioned between water (500 mL)and ether (250 mL), and the layers were separated. The aqueous layer waswashed with ether (2×250 mL) and acidified to pH 1 with concentratedhydrochloric acid. The resulting mixture was cooled in an ice bath andfiltered. The solid was washed with water and dried to afford the titlecompound (25.92 g, 67%) as an off-white solid: ¹ H NMR (400 MHz, CDCl₃)δ 7.90-8.02 (m, 2H), 7.09 (t, J=8.9 Hz, 1H), 2.34 (d, J=1.7 Hz, 3H); IR(chloroform) 3400-2300 (broad), 1694 cm⁻¹ ; MS(ES) m/e 155 [M+H⁺ ; m/e153 [M-H]⁻.

b) t-butyl 4-fluoro-3-methylbenzoate

Isobutylene was bubbled into a suspension of the compound of Example28(a) (3.08 g, 20 mmol) in anhydrous ether (15 mL) in a pressure bottleat -78° C. to give a reaction volume of 50 mL. Trifluoromethanesulfonicacid (0.09 mL, 1 mmol) was added dropwise, and the vessel was tightlysealed and warmed to RT. After 4 h, the reaction was cooled and thevessel was opened. Aqueous 5% sodium bicarbonate was added, and themixture was stirred in a warm water bath to remove the excessisobutylene. Ether was added and the layers were separated. The etherlayer was washed with 5% sodium bicarbonate and with brine, and dried(MgSO₄). Concentration gave the title compound (3.83 g, 91%) as a yellowoil which was used without further purification: TLC (toluene) R_(f)0.73; ¹ H NMR (250 MHz, CDCl₃) δ 7.77-7.89 (m, 2H), 7.01 (t, J=8.9 Hz,1H), 2.30 (d, J=1.8 Hz, 3H), 1.59 (s, 9H); IR (CCl₄) 1715, 1369, 1296,1258, 1167, 1124, 1117, 1105 cm⁻¹ ; MS(ES) m/e 443 [2M+Na]⁺, 421[2M+H]⁺, 211 [M+H]⁺, 155 [M+H-C₄ H₈ ]⁺.

c) t-butyl 4-fluoro-3-(methylaminomethyl)benzoate

A mixture of the compound of Example 28(b) (3.83 g, 18.22 mmol),N-bromosuccinimide (3.57 g, 20.24 mmol), benzoyl peroxide (0.22 g, 0.91mmol), and carbon tetrachloride (90 mL) was heated at reflux. After 16h, the reaction was cooled in an ice bath, filtered, and the filtratewas concentrated. The residue was passed through a short pad of silicagel which was then washed with 20% ethyl acetate:hexane, and thefiltrate was concentrated. The residue was dissolved in THF (90 mL), and40% aqueous methylamine (7.9 mL, 91.1 mmol) was added rapidly. Thereaction was stirred overnight and concentrated. The residue was dilutedwith ether and washed sequentially with 1.0 N sodium hydroxide, water,and brine. The organic phase was dried (MgSO₄), concentrated, and theresidue was chromatographed (silica gel, 10% methanol in 1:1 ethylacetate:chloroform) to give the title compound (2.58 g, 59%) as a yellowoil: TLC R_(f) 0.49 (silica gel, 10% methanol in 1:1 ethylacetate/chloroform); ¹ H NMR (250 MHz, CDCl₃) δ 7.97 (dd, J=7.3, 2.3 Hz,1H), 7.90 (ddd, J=8.4, 5.2, 2.3 Hz, 1H), 7.07 (app t, 1H), 3.83 (s, 2H),2.46 (s, 3H), 1.59 (s, 9H); IR (CCl₄) 1714, 1368, 1297, 1248, 1164, 1106cm⁻¹ ; MS(ES) m/e 240 [M+H]⁺, 184 [M+H-C₄ H₈ ]⁺.

d) t-butyl(S)-4-fluoro-3-[[[4-methoxy-1,4-dioxo-2-[[benzyloxycarbonyl]amino]butyl]methylamino]methyl]benzoate

Dicyclohexylcarbodiimide (453.9 mg, 2.2 mmol) was added to a solution ofthe compound of Example 28(c) (478.6 mg, 2 mmol), N-Cbz-L-aspartic acidβ-methyl ester [J. Am. Chem. Soc., 79, 5697 (1957); J. Am. Chem. Soc.,85, 2483 (1963)] (618.8 mg, 2.2 mmol), and 1-hydroxybenzotriazolehydrate (297.3 mg, 2.2 mmol) in anhydrous dimethylformamide (5 mL) atRT. After 24 h, the mixture was diluted with ether (25 mL) and filtered.The filtrate was concentrated, and the residue was diluted with ether(50 mL) and washed with water (2×10 mL) and brine (10 mL). Drying(MgSO₄), concentration, and chromatography (silica gel, 2:1 hexane:ethylacetate) gave the title compound (0.87 g, 87%) as a colorless oil: TLCR_(f) 0.44 (silica gel, 2:1 hexane:ethyl acetate); ¹ H NMR (250 MHz,CDCl₃) mixture of amide rotamers; δ 7.75-8.00 (m, 2H), 7.15-7.45 (m,5H), 7.00-7.15 (m, 1H), 5.72-5.92 (m, 1H), 4.91-5.25 (m, 3H), 4.45-4.63(m, 2H), 3.66 (s, 3H), 3.13 and 2.89 (2× s, 3H), 2.58-2.94 (m, 2H), 1.56(s,9H); IR (CCl₄) 3415, 3260, 1736, 1716, 1652, 1497, 1368, 1296, 1252,1207, 1163, 1117, 1109 cm⁻¹ ; MS(ES) m/e 1027 [2M+Na]⁺, 1005 [2M+H]⁺,503 [M+H]⁺, 447 [M+H-C₄ H₈ ]⁺.

e) t-butyl(S)-4-fluoro-3-[[[4-methoxy-1,4-dioxo-2-aminobutyl]methylamino]methyl]benzoate

A mixture of the compound of Example 28(d) (0.87 g, 1.73 mmol), 10% Pd/C(184 mg, 0.17 mmol), and methanol (17 mL) was shaken at RT underhydrogen (50 psi). After 1.5 h, the reaction was filtered throughCelite® and concentrated. Chromatography (silica gel, 10% methanol in1:1 ethyl acetate:chloroform) gave the title compound (579.8 mg, 91%) asa colorless oil: TLC R_(f) 0.50 (silica gel, 10% methanol in 1:1 ethylacetate:chloroform); ¹ H NMR (250 MHz, CDCl₃) mixture of amide rotamers;δ 7.80-8.05 (m, 2H), 7.02-7.18 (m, 1H), 4.55-4.88 (m, 2H), 4.15-4.28 (m,1H), 3.71 and 3.70 (2× s, 3H), 3.11 and 2.95 (2× s, 3H), 2.76 (dd,J=16.3, 5.4 Hz, 1H), 2.48-2.67 (m, 1H), 1.74 and 1.57 (2× s, 9H); IR(CCl₄) 3380, 1735, 1717, 1653, 1370, 1298, 1253, 1164, 1112 cm⁻¹ ;MS(ES) m/e 369 [M+H]⁺, 313 [M+H-C₄ H₈ ]⁺.

f) methyl(S)-(-)-2,3,4,5-tetrahydro-7-(t-butoxycarbonyl)-4-methyl-3-oxo-1H-1,4-benzodiazepine-2-acetate

A solution of the compound of Example 28(f) (416.1 mg, 1.13 mmol) and2,6-di-t-butylpyridine (0.51 mL, 2.26 mmol) in anhydrous dimethylsulfoxide (5.7 mL) was heated under argon in an oil bath at 120-125° C.After 17.5 h, the reaction was cooled in ice-water and diluted withwater. The mixture was extracted with ethyl acetate, and the combinedethyl acetate layers were washed with water and brine. Drying (MgSO₄),concentration, and chromatography (silica gel, 3:2 ethyl acetate:hexane)gave the title compound (221.8 mg, 56%) as a nearly colorless solid: TLCR_(f) 0.41 (3:2 ethyl acetate/hexane); [α]_(D) -202.7° (c 1.0,CH₃ OH); ¹H NMR (400 MHz, CDCl₃) δ 7.68 (dd, J=8.5, 1.8 Hz, 1H), 7.59 (d, J=1.8Hz, 1H), 6.50 (d, J=8.5 Hz, 1H), 5.44 (d, J=16.4 Hz, 1H), 5.08-5.16 (m,1H), 4.54 (br d, J=4.5 Hz, 1H), 3.76 (d, J=16.4 Hz, 1H), 3.75 (s, 3H),3.08 (s,3H), 3.00 (dd, J=16.0, 6.9 Hz, 1H), 2.67 (dd, J=16.0, 6.3 Hz,1H), 1.57 (s, 9H); IR (chloroform) 3560-3240 (br), 1731, 1695, 1663,1610, 1369, 1302, 1252, 1171, 1146, 1134 cm⁻¹ ; MS(ES) m/e 719 [2M+Na]⁺,697 [2M+H]⁺, 349 [M+H]⁺.

g) methyl(S)-(-)-7-carboxy-2,3,4,5-tetrahydro-4-methyl-3-oxo-1H-1,4-benzodiazepine-2-acetate

A solution of the compound of Example 28(f) (176.4 mg, 0.506 mmol) inanhydrous dichloromethane (2.5 mL) was cooled to 0° C. and deoxygenatedthrough three evacuation/argon flush cycles. TFA (2.5 mL) was addeddropwise over 3 min, and the resulting solution was stirred for 2 h,during which time the temperature rose to +15° C. The reaction wasconcentrated (20° C.) and the residue was reconcentrated from toluene(35° C.) to remove any residual TFA and give the title compound.

h) methyl(S)-(-)-7-[1-[1'-(t-butoxycarbonyl)-4,4'-bipiperidinyl]carbonyl]-2,3,4,5-tetrahydro-4-methyl-3-oxo-1H-1,4-benzodiazepine-2-acetate

The compound of Example 28(g), 1-hydroxybenzotriazole hydrate (82 mg,0.607 mmol), and 4-(t-butoxycarbonyl)-4,4'-bipiperidine (163 mg, 0.607mmol) were dissolved in anhydrous dimethylformamide (2.5 mL), and thesolution was cooled to 0° C. under argon. Diisopropylethylamine (0.18mL, 1.01 mmol) was added, followed by1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride (116 mg,0.607 mmol), and the reaction was warmed to RT. After 21 h, the reactionwas concentrated and the residue was partitioned between water andchloroform. The layers were separated and the aqueous layer wasextracted with chloroform. Drying (MgSO₄), concentration, andchromatography (silica gel, 10% methanol in 1:1 ethylacetate:chloroform) gave the title compound (215.4 mg, 78%) as a yellowfoam: TLC R_(f) 0.50 (10% methanol in 1:1 ethyl acetate:chloroform);[α]_(D) -140.2° (c 1.0,CH₃ OH); ¹ H NMR (400 MHz, CDCl₃) δ 7.08-7.15 (m,2H), 6.52 (d, J=8.3 Hz, 1H), 5.44 (dd, J=16.4 Hz, 1H), 5.06 (t, J=6.6Hz, 1H), 3.90-4.80 (br m, 4H), 3.75 (s, 3H), 3.73 (d, J=16.4 Hz, 1H),3.08 (s, 3H), 2.99 (dd, J=15.8, 6.6 Hz, 1H), 2.56-2.93 (m, 5H),1.60-1.83 (m, 4H), 1.46 (s, 9H), 1.08-1.42 (m, 6H); IR (CCl₄) 3290,1735, 1689, 1669, 1610, 1436, 1426, 1365, 1286, 1173 cm⁻¹ ; MS(ES) m/e1085 (2M+H)⁺, 565 [M+Na]⁺, 543 [M+H]⁺, 502, 487 [M+H-C₄ H₈ ]⁺.

i)(S)-(-)-7-[1-[1'-(t-butoxycarbonyl)-4,4'-bipiperidinyl]-carbonyl]-2,3,4,5-tetrahydro-4-methyl-3-oxo-1H-1,4-benzodiazepine-2-aceticacid

Using the procedure of Example 27(b)-(c), the compound of Example 28(h)was converted to the title compound.

Example 29 Preparation of(S)-7-[(4,4'-bipiperidin-1-yl)carbonyl]-2,3,4,5-tetrahydro-4-methyl-3-oxo-1H-1,4-benzodiazepine-2-aceticacid

a) 4-fluoro-3-cyanobenzoic acid

To a mixture of 2-fluoro-5-methylbenzonitrile (13.5 g, 0.1 mol),tetrabutylammonium bromide (1.61 g, 5 mmol), and ruthenium(III)chloridetrihydrate (0.261 g, 1 mmol) in dichloromethane (150 mL) was added asolution of sodium hypochlorite (645 mL, 0.45 mol) which had beenneutralized to pH 9 with 20% sulfuric acid. The reaction mixture wasstirred and maintained at pH 9 by adding 20% sodium hydroxide solutionat RT. After 3 h, the aqueous phase was separated and neutralized to pH2 with 20% sulfuric acid and the solid which formed was filtered to givethe title compound (3.4 g, 20%). mp 177-8° C. ¹ H NMR (CDCl₃) δ 7.35 (t,J=8.70 Hz, 1H), 8.38 (m, 1H), 8.42 (m, 1H).

b) t-butyl 4-fluoro-3-cyanobenzoate

To a stirred solution of the compound of Example 29(a) (100 mg, 0.6mmol) in toluene (20 mL) at 80° C. was added a solution ofN,N-dimethylformamide di-t-butyl acetal (1 mL, 4 mmol) in toluene (2 mL)over 10 min. The reaction mixture was heated for 1 h, cooled, washedwith saturated sodium bicarbonate, dried (MgSO₄), and concentrated togive the title compound (70 mg, 52%). MS(ES) m/e 222 [M+H]⁺. ¹ H NMR(CDCl₃) δ 1.60 (9H, s), 7.29 (t, J=8.62 Hz, 1H) 8.25 (m, 1H), 8.28 (m,1H).

c) N-[[2-cyano-4-t-butoxycarbonyl]phenyl]-L-aspartic acid β-methyl ester

A mixture of the compound of Example 29(b) (6 g, 27 mmol), L-asparticacid β-methyl ester hydrochloride (6.1 g, 33 mmol), and sodiumbicarbonate (8.37 g, 100 mmol) in water (20 mL) and dimethyl sulfoxide(80 mL) was heated at 72° C. for 24 h. The reaction mixture was dilutedwith ice cold dilute hydrochloric acid and extracted with ethyl acetate.The combined organic phase was extracted with cold saturated aqueoussodium bicarbonate. The basic extracts were combined, acidified to pH 2with cold dilute hydrochloric acid, and extracted with ethyl acetate.The combined organic extract was dried (MgSO₄), and concentrated to givethe title compound (7.1 g, 75%), MS(ES) m/e 349 [M+H]⁺ ; ¹ H NMR (CDCl₃)δ 1.57 (s, 9H), 3.01 (m, 2H), 3.75 (s, 1H), 4.64 (m, 1H), 5.77 (d,J=8.49 Hz, 1H), 6.71 (d, J=8.93 Hz, 1H), 8.02 (dd, J=8.93, 1.85 Hz, 1H),8.08 (d, J=1.89 Hz, 1H).

d) N-[[2-formyl-4-t-butoxycarbonyl]phenyl]-L-aspartic acid β-methylester

The compound of Example 29(c) (7.0 g, 20 mmol) was dissolved in aceticacid (65 mL) and placed in a hydrogenation bottle. After cooling, wetRaney® nickel (14 g), water (65 mL) and pyridine (65 mL) were added. Theresulting mixture was purged of oxygen with argon for 0.5 h, and shakenin a hydrogen atmosphere (50 psi) at 60° C. for 5 h. The mixture wasdegassed, filtered and the filtrate was azeotroped with toluene at 50°C. under high vacuum. The residual oil was taken up in ethyl acetate(300 mL) and washed with cold dilute hydrochoric acid, dried (MgSO₄),and concentrated to give the title compound as light orange amorphoussolid (5.9 g, 84%). MS(ES) m/e 352 [M+H]⁺ ; 1H NMR (CDCl₃) δ 1.59 (s,9H), 3.00 (m, 2H), 3.74 (s, 1H), 4.77 (m, 1H), 6.73 (d, J=8.88 Hz, 1H),8.03 (d, J=8.85 Hz, 1H), 8.20 (d, J=2.12 Hz, 1H), 9.20 (d, J=8.31 Hz,1H), 9.88 (s, 1H).

e) N-[[2-(N-methyl)aminomethyl-4-t-butoxycarbonyl]phenyl]-L-asparticacid β-methyl ester

A solution of the compound of Example 29(d) (5.9 g, 16.7 mmol) in ethylacetate (100 mL) in a hydrogenation bottle was purged of oxygen withargon for 0.5 h. Platinum oxide (0.6 g), anhydrous MgSO₄ (5.9 g, 49mmol), and a solution of methylamine (3.2M in methanol, 150 mL, 480mmol) were added. The resulting mixture was shaken in a hydrogenatmosphere (50 psi) for 5 h and let stand for 18 h at RT. After additionof acetic acid (75 mL) with cooling, the mixture was again shaken in ahydrogen atmosphere (50 psi) for 2 h, and filtered. The filtrate wasconcentrated and azeotroped with toluene and the residue was desaltedthrough an XAD-2 (350 g) column eluted with water (1.5 L) followed by a50% acetonitrile:water. Two fractions (300 mL) were collected,concentrated and the residue was freeze-dried to give an off-whitepowder. The powder was dissolved in dichloromethane (200 mL),concentrated and the residue was azeotroped with toluene, and dried togive the title compound (5.2 g, 85%). MS(ES) m/e 367 [M+H]⁺.

f) methyl(S)-7-(t-butoxycarbonyl)-2,3,4,5-tetrahydro-3-oxo-4-methyl-1H-1,4-benzodiazepine-2-acetate

To a solution of the compound of Example 29(e) (5.2 g, 14.2 mmol) andtriethylamine (2.0 mL, 14.4 mmol) in dichloromethane (250 mL) at RTunder an argon atmosphere was addedbenzotriazol-1-yloxy-tris(dimethylamino)phosphonium hexafluorophosphate(6.5 g, 14.7 mmol). The reaction mixture was stirred overnight at RT,and washed with an ice-cold solution of dilute hydrochoric acid, water,5% sodium bicarbonate, saturated brine, and then dried (MgSO₄). Thesolution was filtered, concentrated and the residue was chromatographed(silica gel, 40% ethyl acetate:hexane) to give the title compound (1.1g, 22%). MS(ES) m/e 349.2 [M+H]⁺ ; ¹ H NMR (CDCl₃) δ 1.57 (s, 9H), 2.67(dd, J=15.93, 6.30 Hz, 2H), 3.00 (dd, J=15.93, 6.77 Hz, 2H), 3.08 (s,3H), 3.75 (s, 1H), 3.76 (d, J=16.44 Hz, 1H) 4.51 (d, J=4.61, 1H), 5.11(m, 1H) 5.45 (d, J=16.44 Hz, 1H), 6.50 (d, J=8.44 Hz, 1H), 7.55 (d,J=1.88 Hz, 1H), 8.20 (dd, J=8.48, 1.88 Hz, 1H); [α]_(D) -225.1° (c 1,CH₃OH).

g) methyl(S)-7-carboxy-2,3,4,5-tetrahydro-3-oxo-4-methyl-1H-1,4-benzodiazepine-2-acetate

To a solution of the compound of Example 29(f) (160 mg, 0.46 mmol) indichloromethane (5 mL) was added 4M hydrogen chloride:dioxane (5 mL, 20mmol) at RT under argon. The reaction mixture was stirred for 18 h. Thesuspension was concentrated to give the title compound as an off-whitesolid.

h) methyl(S)-7-[(4-t-butoxycarbonyl-4'-bipiperidin)-1-ylcarbonyl]-2,3,4,5-tetrahydro-4-methyl-3-oxo-1H-1,4-benzodiazepine-2-acetate

The compound of Example 29(g), 1-hydroxybenzotriazole (75 mg, 0.55mmol), and 4-t-butoxycarbonyl-4'-bipiperidine were dissolved inanhydrous dimethylformamide (2.5 mL). Diisopropylethylamine (0.16 mL,0.92 mmol) was added, followed by1-(dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride (105 mg, 0.55mmol), and the reaction mixture was stirred overnight at RT under argon.The reaction mixture was partitioned between ice water (50 mL) and ethylacetate (30 mL). The layers were separated and the aqueous phase wasextracted with ethyl acetate (2×30 mL). The combined organic extract wasdried (MgSO₄), concentrated and the residue was chromatographed (silicagel, 2% methanol:dichloromethane) to give the title compound (162 mg,59%). MS(ES) m/e 599.0 [M+H]⁺ ; ¹ H NMR (CDCl₃) δ 1.45 (s, 9H), 1.20 (m,6H), 1.70 (m, 4H), 2.66 (dd, J=16.08, 6.04 Hz, 2H), 2.81 (br s, 4H),2.98 (dd, J=16.08, 7.31 Hz, 2H), 3.06 (s, 3H), 3.71 (d, J=16.44 Hz, 1H),3.73 (s, 1H), 3.76 (d, J=16.44 Hz, 1H) 4.13 (br s, 4H), 5.07 (m, 1H)5.45 (d, J=16.44 Hz, 1H), 6.52 (d, J=8.51 Hz, 1H), 7.09 (m, 2H); [α]_(D)-139.3° (c 1,CH₃ OH).

i)(S)-7-[1-[1'-(t-butoxycarbonyl)-4,4'-bipiperidinyl]carbonyl]-2,3,4,5-tetrahydro-4-methyl-3-oxo-1H-1,4-benzodiazepine-2-aceticacid

Using the procedure of Example 27(b-c), the compound of Example 29(h)was converted to the title compound.

Example 30 Preparation of(S)-7-[[4,4'-bipiperidin-1-yl]carbonyl]-2,3,4,5-tetrahydro-4-methyl-3-oxo-1H-1,4-benzodiazepine-2-aceticacid

a)(S)-7-[[1'-(t-butoxycarbonyl)-4,4'-bipiperidin-1-yl]carbonyl]-2,3,4,5-tetrahydro-4-methyl-3-oxo-1H-1,4-benzodiazepine-2-aceticacid

The compound of Example 27(a) (2.85 g, 5.3 mmol) was taken up in 1:1methanol:THF (30 mL), purged with argon, cooled and treated with 1Nsodium hydroxide (6.3 mL, 6.3 mmol). The resulting solution was stirredat RT for 16 h under argon. The solution was concentrated, diluted withwater (10 mL), and adjusted to pH 5 with 1M acetic acid (litmus paper).The solution was filtered and the solid was dried in vacuo to give titlecompound (2.46 g, 87%).

b)(S)-7-[[4,4'-bipiperidin]-1-ylcarbonyl]-2,3,4,5-tetrahydro-4-methyl-3-oxo-1H-1,4-benzodiazepine-2-aceticacid

The compound of Example 30(a) (1.46 g, 2.77 mmol) was treated with 4Mhydrogen chloride in dioxane (30 mL) for 1 h under argon. The solutionwas concentrated and the residue was taken up in water and adjusted topH 9 (litmus paper) with concentrated ammonium hydroxide and theresulting solution was loaded onto an XAD-2 column (3×7.5 cm). Thecolumn was eluted with water followed by 50% acetonitrile:water.Fractions containing the product were combined and lyophilized to givethe title compound (910 mg, 77%) which was recrystallized from aqueousacetone. mp (shrinking 193) 218-220° C.; ¹ H NMR (400 MHz, DMSO_(d6)) δ1.0-3.9 (m, 26H), 6.5-8.5 (m, 5H); MS (ES) m/e 429.2 [M+H]⁺ ; HPLC k'5.37 (VyDac C18, 4.5×250 mm, gradient, A:acetonitrile B:water-0.1% TFA,5%-60% acetonitrile in 20 min, UV detection at 220 nm); [α]_(D) -180.3°(c 1, MeOH); Anal. (C₂₃ H₃₂ N₄ O₄.2.33 H₂ O) calcd: C, 58.71; H, 7.81;N, 11.91. found: C, 58.67; H, 7.79; N, 11.80.

Example 31 Preparation of(R,S)-7-[(4,4'-bipiperidin-1-yl)carbonyl]-2,3,4,5-tetrahydro-4-methyl-3-oxo-1H-1,4-benzodiazepine-2-aceticacid

Using the procedure of Example 30, except substituting the compound ofExample 8(j) for the compound of Example 27(a), gave the title compoundas a crystalline solid. mp (shrinking 193° C.) 219-220° C.; MS(ES) m/e429 [M+H]⁺ ; HPLC k' 6.5 (Ultrasphere ODS, 4.5×250 mm, gradient,A:acetonitrile B:water-0.1% trifluroracetic acid, 5-60% acetonitriledurings 20 min, UV detection at 220 nm).

Example 32 Preparation of Sodium(R,S)-7-[(4,4'-bipiperidin-1-yl)carbonyl]-2,3,4,5-tetrahydro-4-methyl-3-oxo-1H-1,4-benzodiazepine-2-acetate

The compound of Example 31 (10 mg, 0.023 mmol) was taken up in water (1mL) and 1M sodium hydroxide (0.024 mL, 0.024 mmol) was added. Theresulting solution was lyophilized and the powder was crystallized fromaqueous acetone to give the title compound as a crystalline solid (2mg). mp 220-223° C. [shrinking 204° C.].

Example 33-34

a) The compound of Example 1(a) was separated by chiral HPLC to give thefollowing compounds: 33(a). methyl(R)-8-[[[2-[N-(benzyloxycarbonyl)-4-piperidinyl]ethyl]methylamino]carbonyl]-2,3,4,5-tetrahydro-4-methyl-3-oxo-1H-1,4-benzodiazepine-2-acetate:HPLC t_(R) 30.4 min (Chiralpak AS, 4.6×250 mm, 8 mL/min, 1:1hexane:ethanol-0.5% diethylamine, UV detection at 325 nm); HPLC t_(R)25.8 min (Ultrasphere C18, 4.6×250 mm, gradient, A: acetonitrile B:water-0.1% TFA, 5-60% acetonitrile over 20 min, UV detection at 220 nm)34(a) methyl(S)-8-[[[2-[N-(benzyloxycarbonyl)-4-piperidinyl]ethyl]methylamino]carbonyl]-2,3,4,5-tetrahydro-4-methyl-3-oxo-1H-1,4-benzodiazepine-2-acetate:HPLC t_(R) 22.1 min (Chiralpak AS, 21.2×250 mm, 8 mL/min, 1:1hexane:ethanol-0.5% diethylamine, UV detection at 254 nm); HPLC t_(R)25.9 min (Ultrasphere C18, 4.6×250 mm, gradient, A: acetonitrile B:water-0.1% TFA, 5-60% acetonitrile over 20 min, UV detection at 325 nm).

b) The compounds of Examples 33(a) and 34(a) (100 mg) were separatelydissolved in methanol (15 mL) containing acetic acid (20 drops) weretreated with 10% Pd/C (50 mg) and shaken in a hydrogen atmosphere for 5h. The mixture was filtered through Celite® and concentrated to give thefollowing compounds: 33(b). methyl(R)-2,3,4,5-tetrahydro-4-methyl-3-oxo-8-[[[2-(4-piperidinyl)ethyl]methylamino]carbonyl]-1H-1,4-benzodiazepine-2-acetate.HPLC t_(R) 15.6 min (Ultrasphere C 18, 4.6×250 mm, gradient, A:acetonitrile B: water-0.1% TFA, 5-60% acetonitrile over 20 min, UVdetection at 220 nm) 34(b). methyl(S)-2,3,4,5-tetrahydro-4-methyl-3-oxo-8-[[[2-(4-piperidinyl)ethyl]methylamino]carbonyl]-1H-1,4-benzodiazepine-2-acetate.HPLC t_(R) 15.5 min (Ultrasphere C18, 4.6×250 mm, gradient, A:acetonitrile B: water-0.1% TFA, 5-60% acetonitrile over 20 min, UVdetection at 220 nm).

c) Using the procedure of Example 1(c), except sub-stituting thecompound of Examples 33(b) and 34(b) for the compound of Example 1(b)yielded the following compounds: 33(c).(R)-2,3,4,5-tetrahydro-4-methyl-3-oxo-8-[[[2-(4-piperidinyl)ethyl]methylamino]carbonyl]-1H-1,4-benzodiazepine-2-aceticacid: ¹ H NMR (DMSO-d₆) δ 6.4-8.6 (m, 5H), 5.0-5.6 (m, 2H), 1.0-3.9 (m,22H); [α]_(D) +82.3 (c 0.3, CH₃ OH); MS(ES) m/e 403 [M+H]⁺, [M-H]⁻ 401;HPLC k' 5.0 HPLC t_(R) 15.6 min (Ultrasphere C18, 4.6×250 mm, gradient,A: acetonitrile B: water-0.1% TFA, 5-60% acetonitrile over 20 min, UVdetection at 220 nm); Anal. (C₂₁ H₃₀ N₄ O₄.2.25 C₂ HF3O₂.2.75 H₂ O)calcd: C, 50.44; H, 5.78; N, 9.94. found: C, 50.42; H, 5.89; N, 10.1734(c).(S)-2,3,4,5-tetrahydro-4-methyl-3-oxo-8-[[[2-(4-piperidinyl)ethyl]methylamino]carbonyl]-1H-1,4-benzodiazepine-2-aceticacid: ¹ H NMR (DMSO-d₆) δ 6.4-8.6 (m, 5H), 5.0-5.6 (m, 2H), 1.0-3.9 (m,22H); [α]_(D) -110 (c 0.45, CH₃ OH); MS(ES) m/e 403 [M+H]⁺, [M-H]⁻ 401;HPLC k' 5.0 HPLC t_(R) 15.6 min (Ultrasphere C18, 4.6×250 mm, gradient,A: acetonitrile B: water-0.1% TFA, 5-60% acetonitrile over 20 min, UVdetection at 220 nm); Anal. (C₂₁ H₃₀ N₄ O₄.1.5 C₂ HF₃ O₂.1.25 H₂ O)calcd: C, 43.22; H, 5.37; N, 7.91. found: C, 43.10; H, 5.17; N, 8.11.

Example 35 Preparation of(S)-2,3,4,5-tetrahydro-4-methyl-3-oxo-8-[[[2-(4-piperidinyl)ethyl]methylamino]carbonyl]-1H-1,4-benzodiazepine-2-acetic acid

a) t-butyl(S)-4-[N-(t-butoxycarbonyl)-(methylamino)-methyl]-3-[2-(1,4-dimethoxy-1,4-dioxo-2-butyl)amino]benzoate

A solution of t-butyl3-amino-4-[N-(t-butoxycarbonyl)-N-(methyl)-aminomethyl]benzoate (10.4 g,0.03 mol) in dichloromethane (75 mL) was treated with a solution ofdimethyl (R)-2-(trifluoromethylsulfonyloxy)-succinate [Liebigs Ann.Chem., 314-333 (1986)] (8.8 g, 0.03 mol) and 2,6-di-t-butylpyridine (6g, 0.03 mol) in dichloromethane (10 mL) and the mixture was stirred for4 d. The mixture was washed with dilute hydrochoric acid, and theorganic phase was washed with aqueous sodium carbonate, dried (MgSO₄)and concentrated. The residue was chromatographed (silica gel, 10% ethylacetate:hexane) and concentrated to give the title compound (73%).

b) t-butyl(S)-4-(methylamino)methyl-3-[2-(1,4-dimethoxy-1,4-dioxo-2-butyl)amino]benzoate

A solution of the compound of Example 35(a) (13 g, 0.027 mol) indichloromethane (25 mL) was treated with 4M hydrogen chloride in dioxane(60 mL), stirred for 45 min, and washed with 5% sodium carbonate. Theorganic phase was dried (MgSO₄) and concentrated to give the titlecompound (6.5 g).

c) methyl(S)-8-(t-butoxycarbonyl)-2,3,4,5-tetrahydro-4-methyl-3-oxo-1H-1,4-benzodiazepine-2-acetate

A solution of the compound of Example 35(b) (6.5 g, 0.015 mol) in xylene(100 mL) was heated to 130° C. for 14.5 h, cooled, and concentrated. Theresidue was stirred with ether, filtered, and concentrated. The residuewas chromatographed (silica gel, 0.5-0.75% methanol: dichloromethane)and fractions containing product were pooled, concentrated, diluted withether and stored in the cold for 3 days. The mixture was filtered andthe filtrate was concentrated to give the title compound (2.7 g, 52%).

d) methyl(S)-8-carboxy-2,3,4,5-tetrahydro-4-methyl-3-oxo-1H-1,4-benzodiazepine-2-acetatehydrochloride

The compound of Example 35(c) (2.8 g) in dichloromethane (10 mL) wastreated with 4M hydrogen chloride in dioxane (25 mL) and stirred for 16h. The mixture was concentrated to give the title compound.

e) methyl(S)-2,3,4,5-tetrahydro-4-methyl-3-oxo-8-[[[2-[N-trifluoroacetyl-4-piperidinyl]ethyl]methylamino]carbonyl]-1H-1,4-benzodiazepine-2-acetate

The compound of Example 35(d) (310 mg, 1.1 mmol) anddiisopropylethylamine (285 mg, 2.2 mmol) in dimethylformamide (2.5 mL)was added to a mixture ofN-methyl-2-(N-trifluoroacetyl-4-piperidinyl)ethylamine hydrochloride(300 mg, 1.1 mmol), diisopropylethylamine (140 mg, 1.1 mmol), and1-hydroxybenzotriazole hydrate (150 mg, 1.1 mmol) in dimethylformamide(2.5 mL) followed by 1-(3-dimethylaminopropyl)-3-ethylcarbodiimidehydrochloride (220 mg, 1.1 mmol). The mixture was stirred for 16 h,concentrated and the residue was triturated with ethyl acetate andwater. The combined organic phase was washed with brine, dried (MgSO₄)and concentrated. The residue was chromatographed (silica gel, 0.5-1.5%methanol:dichloromethane) and fractions containing the product werecombined and concentrated to give the title compound (260 mg, 51%).[α]_(D) -75.6 (c 1,CH₃ OH); MS(ES) m/e 513 [M+H]⁺ ; 557 [M+HCO₂ ⁻ ]⁻ ;HPLC t_(R) 8.22 min (WHELK-O, 4.6×25 mm, 1 mL min, 10% methanol:water,UV detection at 220 nM); Anal. (C₂₄ H₃₁ F₃ N₄ O₅.1.5 H₂ O) calcd: C,53.33; H, 6.34; N, 10.36 found: C, 53.39, H, 6.23; N, 9.98.

f)(S)-2,3,4,5-tetrahydro-4-methyl-3-oxo-8-[[[2-[4-piperidinyl]ethyl]methylamino]carbonyl]-1H-1,4-benzodiazepine-2-aceticacid

The compound of Example 35(e) was stirred with sodium hydroxide inmethanol/water to give the title compound.

Example 36 Preparation of methyl(S)-7-t-butoxycarbonyl-2,3,4,5-tetrahydro-3-oxo-2-(2-phenylethyl)-1H-2-benzazepine-4-acetate

a) methyl5-(t-butoxycarbonyl)-2-[N-(butoxycarbonyl)-N-(2-phenylethyl)aminomethyl]cinnamate

To a stirred solution of the compound of Preparation 8(d) (4.9 g, 10mmol) in dry acetonitrile (15 mL) was added methyl acrylate (3 mL, 40mmol), diisopropylethylamine (3.5 mL, 20 mmol), tri-o-tolylphoshine (304mg, 1 mmol) and palladium acetate (112 mg, 0.5 mmol). The reaction wasrepeatedly evacuated and flushed with argon and then heated to 80° C.After 1 h, an additional amount of tri-o-tolylphoshine (304 mg, 1 mmol)and palladium acetate (112 mg, 0.5 mmol) was added and the reaction wasstirred for 2 h. The reaction was then cooled to RT, concentrated,suspended with ether (100 mL) and petroleum ether (50 mL), filtered freeof insoluble materials and concentrated. The residual oil was purifiedby flash chromatography (silica gel, 15% ethyl acetate:hexane) to givethe title compound (4.91 g, 99%). ¹ H NMR (CDCl₃, 400 MHz) (mixture ofamide rotomers) δ 1.42 and 1.49 (9H, 2 br s), 1.60 (9H, s), 2.74 and2.84 (2H, 2 br s), 3.28 and 3.42 (2H, 2 br s), 3.82 (3H, s), 4.47 and4.57 (2H, 2 br s), 6.43 (1H, d, J=14.8 Hz), 7.05-7.30 (6H, m), 7.82-7.92(1H, m), 7.97 (1H, dd), 8.15 (1H, d, J=1.4 Hz).

b) methyl5-(t-butoxycarbonyl)-2-[N-(butoxycarbonyl)-N-(2-phenylethyl)aminomethyl]phenylpropionate

The compound of Example 36(a) (4.91 g, 9.9 mmol) was hydrogenated at 55psi H₂ over 10% Pd/C (2.5 g) in methanol (100 mL) for 7 h. Afterfiltration through a pad of Celite® and concentration, the remaining oilwas purified by flash chromatography (silica gel, 15% ethylacetate:hexane) to give title compound as a clear oil (4.48 g, 91%). ¹ HNMR (CDCl₃, 400 MHz) (mixture of amide rotomers) δ 1.42 and 1.49 (9H, 2br s), 1.60 (9H, s), 2.58 (2H, t), 2.77 and 2.82 (2H, 2 br s), 2.92 (2H,br s), 3.31 and 3.42 (2H, 2 br s), 3.70 (3H, s), 4.38 and 4.47 (2H, 2 brs), 7.07-7.31 (6H, m), 7.78 (1H, s), 7.81 (1H, dd).

c)5-(t-butoxycarbonyl)-2-[N-(butoxycarbonyl)-N-(2-phenylethyl)aminomethyl]phenylpropionicacid

The compound of Example 36(b) (4.48 g, 9 mmol) was treated with 1Nsodium hydroxide (10 mL) in dioxane (50 mL) for 16 h at RT. Afteracidification with 1N hydrochloric acid (10 mL), the reaction wasextracted with ethyl acetate (2×100 mL), washed with brine, dried(MgSO₄), filtered and concentrated to give the title compound as a whitesolid (4.12 g, 95%). TLC R_(f) 0.5 (silica gel, 95:4:1,chloroform:methanol:acetic acid).

d)5-(t-butoxycarbonyl)-2-[N-(butoxycarbonyl)-N-(2-phenylethyl)aminomethyl]phenylpropionylfluoride

To a stirred solution of the compound of Example 36(c) (4.12 g, 8.5mmol) under argon in dry dichloromethane (25 mL) was added pyridine(0.73 mL, 9 mmol) followed by cyanuric fluoride (0.56 mL, 6.1 mmol).After stirring for 2 h, the reaction became a thick suspension. Thereaction was then concentrated, taken up in ether (100 mL) and, filteredthrough a pad of Celite to remove insoluble materials. The filtrate waswashed with cold water, dried (MgSO₄), filtered and concentrated to givethe title compound as a clear oil (4.21 g, 96%).

e)4(R)-benzyl-3-[3-[5-(t-butoxycarbonyl)-2-[N-(t-butoxycarbonyl)-N-(2-phenylethyl)aminomethyl]phenyl]propionyl]-2-oxazolidinone

To a stirred solution of (R)-4-benzyl-2-oxazolidinone (1.8 g, 10 mmol)in dry THF (25 mL) at -78° C. under argon was added via syringe,dropwise, a solution of 2.5N n-butyllithium in hexane (3.8 mL). Afterstirring for 15 min, a solution of the above the compound of Example36(d) (4.21 g, 8.5 mmol) in THF (10 mL) was added dropwise over 5 min.The reaction was stirred for 1 h at -78° C. and quenched with saturatedaqueous ammonium chloride (50 mL). The reaction was allowed to warm toRT, extracted with ethyl acetate, dried (MgSO₄), filtered andconcentrated. The residue was purified by flash chromatography (silicagel, 25% ethyl acetate:hexane) to give the title compound as a whitesolid (5.13 g, 89%). ¹ H NMR (CDCl₃, 400 MHz) (mixture of amiderotomers) δ 1.42 and 1.49 (9H, 2 br s), 1.59 (9H, s), 2.75 (1H, dd),2.78 (1H, m), 2.87 (1H, m), 3.01 (2H, m), 3.18 (1H, m), 3.23 (1H, dd),3.31 (1H, dd), 3.33 (1H, m), 3.44 (1H, m), 4.18 (2H, m), 4.46 and 4.55(2H, 2 br s), 4.67 (1H, m), 7.16 (6H, m), 7.29 (5H, m), 7.81 (1H, d),7.82 (1H, s).

f)4(R)-benzyl-3-[3-[5-(t-butoxycarbonyl)-2-[N-(t-butoxycarbonyl)-N-(2-phenylethyl)aminomethyl]phenyl]-2(S)-(methoxycarbonylmethyl)propionyl]-2-oxazolidinone

To a stirred solution of the compound of Example 36(e) (5.13 g, 8 mmol)in dry THF (75 mL) with stirring at -78° C. under argon was addeddropwise via syringe a solution of 1N lithium hexamethyldisilazane inTHF (8.2 mL). After stirring for 15 min, methyl bromoacetate (3.75 mL,40 mmol) was added in one portion. The reaction was allowed to warm to-28° C. and stirred for an additional 2 h, quenched with saturatedaqueous ammonium chloride, extracted with ethyl acetate, washed withbrine, dried (MgSO₄), filtered and concentrated. The residue waspurified by column chromatography (silica gel, 20% ethyl acetate:hexane)to give the title compound as a white solid (3.00 g, 53%). HPLC k' 2.9,minor diastereomer k' 6.4, d.e.88% (Ultrasphere, 4.6×250 mm, 1.5 mL/min,20% ethyl acetate:hexane, UV detection at 254 nm); ¹ H NMR (CDCl₃, 400MHz) δ 1.40 and 1.47 (9H, 2 br s), 1.52 (9H, s), 2.44 and 2.52 (1H, br sand brd), 2.70 (1H, dd), 2.80 (2H, m), 2.88 (2H, m), 3.00 (1H, dd), 3.31(1H, dd), 3.39 (2H, m), 3.67 (3H, s), 3.79 (1H, t), 4.00 (1H, d), 4.30and 4.38 (1H, br s and br d), 4.55 (3H, m), 7.10-7.35 (11H, m), 7.72(1H, s), 7.79 (1H, dd).

g)(S)-3-[5-(t-butoxycarbonyl)-2-[N-(t-butoxycarbonyl)-N-(2-phenylethyl)aminomethyl]phenyl]-2-(methoxycarbonylmethyl)-propionicacid hydrochloride

To a stirred solution of the compound of Example 36(f) (1.47 g, 2 mmol)in (3:1) THF:water (16 mL) at 0° C. was added dropwise a solution oflithium hydroperoxide made from 30% hydrogen peroxide (0.7 mL) andlithium hydroxide hydrate (90 mg, 2.1 mmol) in water (2.1 mL). Afterstirring for 1 h at 0° C., excess peroxide was destroyed with a solutionof sodium sulfite (1.28 g) in water (6.2 mL) dropwise with cooling. Thereaction was then acidified with 3N hydrochloric acid and extracted withdichloromethane (2×50 mL), dried (MgSO₄), filtered and concentrated.

h)(S)-3-[5-(t-butoxycarbonyl)-2-[N-(t-butoxycarbonyl)-N-(2-phenylethyl)aminomethyl]phenyl]-2-(methoxycarbonylmethyl)-propionicacid hydrochloride

The compound of Example 36(g) was treated with a solution of 4N hydrogenchloride in dioxane (15 mL) with stirring for 10 min at RT andconcentrated without heating. The residue was taken up in ethyl acetateand concentrated to remove excess hydrogen chloride and crystallizedfrom ethyl acetate:ether to give the title compound (0.75 g, 76%) as awhite solid. ¹ H NMR (d₄ -MeOH, 400 MHz) δ 1.60 (9H, s), 2.68 (1H, dd),2.81 (1H, dd), 2.98-3.18 (5H, m), 3.42 (2H, m), 3.68 (3H, s), 4.40 (1H,d, J=13.6 Hz), 4.51 (1H, d, J=13.6 Hz), 7.32 (5H, m), 7.58 (1H, d, J=8.0Hz), 7.90 (1H, dd), 7.95 (1H, d, J=1.4 Hz).

i) methyl(S)-7-(t-butoxycarbonyl)-2,3,4,5-tetrahydro-3-oxo-2-(2-phenylethyl)-1H-2-benzazepine-4-acetate

To a stirred solution of the compound of Example 36(h) (0.35 g, 0.71mmol) in dry N,N-dimethylformamide (20 mL) at 0° C. in a Dewer flask wasadded triethylamine (200 mL, 1.42 mmol), sodium bicarbonate (300 mg,3.57 mmol), and diphenylphosphoryl azide (250 mL, 1.2 mmol). Afterstirring at 0° C. for 24 h, the reaction was concentrated, taken up inethyl acetate (100 mL), washed with water, brine, dried (MgSO₄),filtered and concentrated. The residue was purified by chromatography(silica gel, 35% ethyl acetate:hexane) to give the title compound as asticky white foam (243 mg, 78%). HPLC k' 2.13, k' 1.28(enantiomer) >99.5% optically pure (Chiralpak AS, 0.46×250 mm, 1 mL/min,20% EtOH:n-hexane, UV detection at 254 nm); [α]_(D) -71.2 (c 1.4, MeOH);¹ H NMR (CDCl₃, 400 MHz) δ 1.60 (9H, s), 2.42 (1H, dd), 2.74 (2H, t),2.89 (1H, dd), 3.02 (2H, m), 3.60 (1H, m), 3.72 (3H, s), 3.81 (1H, d,J=16.7 Hz), 3.81 (1H, m), 5.20 (1H d, J=16.7 Hz), 7.00-7.23 (6H, m),7.69 (1H, s), 7.71 (1H, d).

Example 37 Preparation of(R,S)-8-[[(4,4'-bipiperidin-1-yl]carbonyl]-2,3,4,5-tetrahydro-2-methyl-3-oxo-1H-2-benzazepine-4-aceticacid

a) methyl(R,S)-8-carboxy-2,3,4,5-tetrahydro-2-methyl-3-oxo-1H-2-benzazepine-4-acetate

Using the procedure of Preparation 8(a)-(g), except substituting4-bromo-3-methylbenzoic acid for 3-bromo-4-methylbenzoic acid andsubstituting methylamine for phenethylamine, gave the title compound.

b)(R,S)-8-[[(4,4'-bipiperidin-1-yl)]carbonyl]-2,3,4,5-tetrahydro-2-methyl-3-oxo-1H-2-benzazepine-4-aceticacid

Using the procedures of Example 25(b)-(c), except substituting thecompound of Example 37(a) for the compound of Example 25(a), gave thetitle compound. HPLC k' 3.23 (PRP-1; 15% acetonitrile/water-0.1% TFA); ¹H NMR (400 MHz, CD₃ OD) δ 7.21-7.30 (m, 3H), 5.37 (d, J=16.6 Hz, 1H),4.58-4.74 (m, 1H), 4.07 (d, J=16.6 Hz, 1H), 3.84-3.96 (m, 1H), 3.68-3.84(m, 1H), 3.40 (br d, J=12.6 Hz, 2H), 3.17 (dd, J=17.7, 4.0 Hz, 1H),2.69-3.17 (m, 6H), 3.01 (s, 3H), 2.47 (dd, J=17.0, 4.7 Hz, 1H),1.10-2.10 (m, 10H); MS(ES) m/e 428.2 [M+H]⁺, 214.6, 205.4; Anal. (C₂₄H₃₃ N₃ O₄.2 CF₃ CO₂ H.H₂ O) calcd: C, 49.93; H, 5.54; N, 6.24. found: C,50.17; H, 5.32; N, 6.20.

Example 38 Preparation of(R,S)-2,3,4,5-tetrahydro-7-[1-[4-(4-pyridyl)piperazinyl]carbonyl]-4-methyl-3-oxo-1H-1,4-benzodiazepine-2-aceticacid

Methyl(R,S)-7-carboxy-2,3,4,5-tetrahydro-4-methyl-3-oxo-1H-1,4-benzodiazepine-2-acetate(2.0 mmol) dissolved in DMF (30 mL) was treated with1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride (2.2 mmol),1-HOBT (285 mg, 2.1 mmol) and adjusted to pH 7 with triethylamine. Themixture was treated with 1-(pyrid-4-yl)piperazine (2.4 mmol), stirred atRT for 48 h, concentrated and the residue was purified by flashchromatography to yield the title compound. HPLC k' 4.01 (PRP-1; 12%acetonitrile/water-0.1% TFA); ¹ H NMR (400 MHz, CD₃ OD) δ 8.17 (d, J=7.4Hz, 2H), 7.12-7.24 (m, 4H), 6.63 (d, J=9.0 Hz, 1H), 5.59 (d, J=16.7 Hz,1H), 5.21 (dd, J=9.0, 5.1 Hz, 1H), 3.90 (d, J=16.7 Hz, 1H), 3.72-3.90(m, 8H), 3.04 (s, 3H), 2.94 (dd, J=16.6, 9.0 Hz, 1H), 2.66 (dd, J=16.8,5.1 Hz, 1H); MS(ES) m/e 424.2 [M+H]⁺ ; Anal. (C₂₂ H₂₅ N₅ O₄.2 CF₃ CO₂ H)calcd: C, 47.93; H, 4.18; N, 10.75. found: C, 47.86; H, 4.40; N, 10.87.

Example 39 Preparation of(R,S)-7-[[[4-(aminoiminomethyl)phenyl]-methylamino]carbonyl]-2,3,4,5-tetrahydro-2-[2-(2-thienyl)ethyl]-3-oxo-1H-2-benzazepine-4-aceticacid

Using the procedure of Preparation 8(c)-(j), except substituting2-(2-thienyl)ethanamine for 2-phenethylamine in the procedure ofPreparation 8(c) and substituting sodium borohydride and nickel chloridehexahydrate for Pd/C and hydrogen in the procedure of Preparation 8(f)and substituting triethylamine in refluxing toluene for sodium methoxidein methanol in the procedure of Preparation 8(g) and substitutinglithium hydroxide in aqueous THF for aqueous sodium hydroxide inmethanol in the procedure of Preparation 8(j), gave the title compound.

Example 40 Preparation of(R,S)-2,3,4,5-tetrahydro-3-oxo-4-(2-phenylethyll)-7-[[[3-(4-piperidnyl]propyl]methylamino]-carbonyl]-1H-1,4-benzodiazepine-2-aceticacid

Methyl(R,S)-7-carboxy-4-(2-phenylethyl)-1,3,4,5-tetrahydro-3-oxo-2H-1,4-benzodiazepine-2-acetate(2.0 mmol) dissolved in DMF (30 mL) was treated withbenzotriazol-1-yloxy-tris(dimethylamino)phosphonium hexafluorophosphate(2.2 mmol), 1-HOBT (285 mg, 2.1 mmol) and adjusted to pH 7 withtriethylamine. The mixture was treated withN-methyl-3-[N-(tert-butoxycarbonyl)piperidin-4-yl]]propanamine (2.4mmol), stirred at RT for 48 h, concentrated and the residue was purifiedby flash chromatography.

The product is dissolved in 50% TFA/methlene chloride at RT and stirred1 h. The solution is concentrated by evaporation of the TFA andmethylene chloride, redissolved in methylene chloride andre-concentrated.

A portion of the compound (0.26 mmol) was dissolved in methanol (9 mL),and 1.0 N sodium hydroxide (0.81 mL, 0.81 mmol) was added. The solutionwas stirred at RT overnight, and concentrated. The residue was dissolvedin water/acetonitrile (3 mL), cooled to 0° C., and acidified with TFA(0.21 mL, 2.7 mmol). The solution was concentrated and the residue waspurified by reversed-phase flash chromatography (C-18 silica gel,AN/W-TFA). Concentration and lyophilization gave the title compound.MS(ES) m/e 507 [M+H]⁺, 505 [M-H]⁻.

Example 41 Preparation of(R,S)-2,3,4,5-tetrahydro-4-methyl-3-oxo-7-[[[3-(4-piperidnyl]propyl]methylamino]carbonyl]-1H-1,4-benzodiazepine-2-aceticacid

Using the procedure of Example 40, except substituting the compound ofExample 8(i) for methyl(R,S)-7-carboxy-4-(2-phenylethyl)-1,3,4,5-tetrahydro-3-oxo-2H-1,4-benzodiazepine-2-acetate,gave the title compound.

Example 42 Preparation of(R,S)-2,3,4,5-tetrahydro-4-isopropyl-3-oxo-8-[[[2-(4-piperidinyl)ethyl]methylamino]carbonyl]-1H-1,4-benzodiazepine-2-acetic acid

a) methyl(R,S)-8-carboxy-2,3,4,5-tetrahydro-4-isopropyl-3-oxo-1H-1,4-benzodiazepine-2-acetate

Using the procedure of Example 2(a) except substituting isopropylaminefor isopentylamine and substituting diisopropylethylamine in refluxingxylene for sodium methoxide in methanol in the cyclization step gave thetitle compound.

b) methyl(R,S)-2,3,4,5-tetrahydro-4-isopropyl-3-oxo-8-[[[2-(4-pyridyl)ethyl]methylamino]carbonyl]-1H-1,4-benzodiazepine-2-acetate

Using the procedure of Example 2(b), except substituting the compound ofExample 42(a) for the compound of Example 2(b), gave the title compound.

c) methyl(R,S)-2,3,4,5-tetrahydro-4-isopropyl-3-oxo-8-[[[2-(4-piperidinyl)ethyl]methylamino]carbonyl]-1H-1,4-benzodiazepine-2-acetate

Using the procedure of Example 2(c), except substituting the compound ofExample 42(b) for the compound of Example 2(b), gave the title compound.

d)(R,S)-2,3,4,5-tetrahydro-4-isopropyl-3-oxo-8-[[[2-(4-piperidinyl)ethyl]methylamino]carbonyl]-1H-1,4-benzodiazepine-2-aceticacid

Using the procedure of Example 2(d), except substituting the compound ofExample 42(c) for the compound of Example 2(c), gave the title compound.

Example 43 Preparation of(S)-2,3,4,5-tetrahydro-4-(2-phenylethyl)-3-oxo-8-[[[2-(4-piperidinyl)ethyl]methylamino]carbonyl]-1H-1,4-benzodiazepine-2-aceticacid

Using the procedure of Example 35, except substituting t-butyl3-amino-4-[N-(t-butoxycarbonyl)-N-(2-phenylethyl)aminomethyl]benzoatefor t-butyl3-amino-4-[N-(t-butoxycarbonyl)-N-(methyl)-aminomethyl]benzoate inExample 35(a) gave the title compound.

Example 44 Preparation of(R,S)-7-[[[4-(aminoiminomethyl)phenyl]-carbonyl]methylamino]-2,3,4,5-tetrahydro-3-oxo-2-(2-phenylethyl]-1H-2-benzazepine-4-aceticacid

a) methyl(R,S)-7-(benzyloxycarbonyl)amino-2,3,4,5-tetrahydro-3-oxo-2-(2-phenylethyl]-1H-2-benzazepine-4-acetate

A mixture of methyl(R,S)-7-carboxy-2,3,4,5-tetrahydro-3-oxo-2-(2-phenylethyl)-1H-2-benzazepine-4-acetate(2.67 mmol), triethylamine (0.42 mL, 5.87 mmol) and diphenylphosphorylazide (0.62 mL, 2.80 mmol) in toluene (20 mL) was heated at105° C. for 0.5 h. After the temperature was lowered to 80° C., themixture was treated with benzyl alcohol (0.60 mL, 0.42 mmol), stirredfor 14 h and concentrated. The residue was purified by flashchromatography to give the title compound.

b) methyl(R,S)-7-[N-(benzyloxycarbonyl)methylamino]-2,3,4,5-tetrahydro-3-oxo-2-(2-phenylethyl]-1H-2-benzazepine-4-acetate

The compound of Example 169(a) was treated with sodium hydride andiodomethane in 4:1 THF:dimethylformamide to give the title compound.

c) methyl(R,S)-7-methylamino-2,3,4,5-tetrahydro-3-oxo-2-(2-phenylethyl]-1H-2-benzazepine-4-acetate

Using the procedure of Example 19(c), except substituting the compoundof Example 169(b) for the compound of Example 19(b) gave the titlecompound.

d) methyl(R,S)-7-[[[4-(N-benzyloxycarbonyl)(aminoiminomethyl)phenyl]carbonyl]methylamino]-2,3,4,5-tetrahydro-3-oxo-2-(2-phenylethyl]-1H-2-benzazepine-4-acetate

The compound of Example 169(b) is treated with4-[N-(benzyloxycarbonyl)(aminoininomethyl)]benzoic acid, BOP andtriethylamine in 5:4 dichloromethane:THF to give the title compound.

e) methyl(R,S)-7-[[[4-(aminoimiomethyl)phenyl]carbonyl]-amino]-2,3,4,5-tetrahydra-3-oxo-2-(2-phenylethyl]-1H-2-benzazepine-4-acetate

Using the procedure of Example 19(c)-(d), except substituting thecompound of Example 169(c) for the compound of Example 19(b) gave thetitle compound.

Example 45 Preparation of(R,S)-2,3,4,5-tetrahydro-3-oxo-4-methyl-8-[[[2-[(2-amino)pyrid-4-yl]ethyl]methylamino]carbonyl]-1H-1,4-benzodiazepine-2-aceticacid

Methyl(R,S)-8-carboxy-4-methyl-1,3,4,5-tetrahydro-3-oxo-2H-1,4-benzodiazepine-2-acetate(2.0 mmol) dissolved in DMF (30 mL) was treated withbenzotriazol-1-yloxy-tris(dimethylamino)phosphonium hexafluorophosphate(2.2 mmol), 1-HOBT (285 mg, 2.1 mmol) and adjusted to pH 7 withtriethylamine. The mixture was treated with2-[2-(amino)pyrid-4-yl]-N-(methyl)-ethanamine (2.4 mmol), stirred at RTfor 48 h, concentrated and the residue was purified by flashchromatography.

A portion of the compound (0.25 mmol) was dissolved in methanol (9 mL),and 1.0 N sodium hydroxide (0.81 mL, 0.81 mmol) was added. The solutionwas stirred at RT overnight, and concentrated. The residue was dissolvedin water/acetonitrile (3 mL), cooled to 0° C., and acidified with TFA(0.21 mL, 2.7 mmol). The solution was concentrated and the residue waspurified by reversed-phase flash chromatography (C-18 silica gel,AN/W-TFA).

Example 46

Parenteral Dosage Unit Composition

A preparation which contains 20 mg of the compound of Example 1 as asterile dry powder is prepared as follows: 20 mg of the compound isdissolved in 15 mL of distilled water. The solution is filtered understerile conditions into a 25 mL multi-dose ampoule and lyophilized. Thepowder is reconstituted by addition of 20 mL of 5% dextrose in water(D5W) for intravenous or intramuscular injection. The dosage is therebydetermined by the injection volume. Subsequent dilution may be made byaddition of a metered volume of this dosage unit to another volume ofD5W for injection, or a metered dose may be added to another mechanismfor dispensing the drug, as in a bottle or bag for IV drip infusion orother injection-infusion system.

Example 46

Oral Dosage Unit Composition

A capsule for oral administration is prepared by mixing and milling 50mg of the compound of Example 1 with 75 mg of lactose and 5 mg ofmagnesium stearate. The resulting powder is screened and filled into ahard gelatin capsule.

Example 47

Oral Dosage Unit Composition

A tablet for oral administration is prepared by mixing and granulating20 mg of sucrose, 150 mg of calcium sulfate dihydrate and 50 mg of thecompound of Example 1 with a 10% gelatin solution. The wet granules arescreened, dried, mixed with 10 mg starch, 5 mg talc and 3 mg stearicacid; and compressed into a tablet.

The foregoing is illustrative of the making and using of this invention.This invention, however, is not limited to the precise embodimentsdescribed herein, but encompasses all modifications within the scope ofthe claims which follow.

What is claimed is:
 1. A compound whichis:(R,S)-7-[(4,4'-bipiperidin-1-yl)carbonyl]-2,3,4,5-tetrahydro-4-methyl-3-oxo-1H-1,4-benzodiazepine-2-aceticacid; or(S)-7-[(4,4'-bipiperidin-1-yl)carbonyl]-2,3,4,5-tetrahydro-4-methyl-3-oxo-1H-1,4-benzodiazepine-2-aceticacid, or a pharmaceutically acceptable salt thereof.
 2. A pharmaceuticalcomposition comprising a compound according to claim 1 and apharmaceutically acceptable carrier.
 3. A method for inhibiting plateletaggregation which comprises administering a compound according to claim1 to a mammal in need thereof.
 4. A method for treating myocardialinfarction, thrombosis, embolism, stroke and infarct-related disorders,or restenosis following angioplasty, which comprises administering acompound according to claim 1 to a mammal in need thereof.
 5. A methodfor inhibiting reocclusion of an artery or vein following thrombolytictherapy comprising administering a compound according to claim 1 and athrombolytic agent.
 6. A compound according to claim 1 which is(S)-7-[(4,4'-bipiperidin-1-yl)carbonyl]-2,3,4,5-tetrahydro-4-methyl-3-oxo-1H-1,4-benzodiazepine-2-aceticacid, or a pharmaceutically acceptable salt thereof.
 7. A pharmaceuticalcomposition comprising a compound according to claim 6 and apharmaceutically acceptable carrier.
 8. A method for treating acutemyocardial infarction, deep vein thrombosis, pulmonary embolism,transient ischemia attack, unstable angina, stroke, infarct-relateddisorders, or restenosis following angioplasty, which comprisesadministering a compound according to claim 6 to a mammal in needthereof.
 9. A pharmaceutical composition according to claim 2 comprisinga 50 mg tablet.
 10. A pharmaceutical composition according to claim 7comprising a 50 mg tablet.
 11. A method for inhibiting plateletaggregation which comprises administering a compound according to claim6 to a mammal in need thereof.
 12. A method for inhibiting reocclusionof an artery or vein following thrombolytic therapy comprisingadministering a compound according to claim 6 and a thrombolytic agent.13. A method for inhibiting platelet aggregation which comprisesadministering a compound according to claim 6, and a cyclooxygenaseinhibitor, thromboxane antagonist, thromboxane synthesis inhibitor,heparin, thrombin inhibitor, ADP receptor inhibitor or ticlopidine. 14.A method for inhibiting platelet aggregation which comprisesadministering a compound according to claim 6 and aspirin.
 15. A methodfor inhibiting platelet aggregation which comprises administering acompound according to claim 6 and clopidogrel.
 16. A process forpreparing(S)-7-[(4,4'-bipiperidin-1-yl)carbonyl]-2,3,4,5-tetrahydro-4-methyl-3-oxo-1H-1,4-benzodiazepine-2-aceticacid, or a pharmaceutically acceptable salt thereof, which comprisestreating a compound of the formula: ##STR6## wherein R is H, C₁₋₆ alkyl,benzyl or a carboxy protecting group;R³ is methyl; X is H; and R^(6') is(4,4'-bipiperidin-1-yl)carbonyl wherein the basic nitrogen isprotected;with a reagent, in any order, (i) to remove an aminoprotecting group from R^(6') ; and, if necessary, (ii) to remove acarboxy protecting group from CO₂ R; and (iii) form a pharmaceuticallyacceptable salt thereof.